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鸡传染性喉气管炎病毒WF03株关键抗原基因和毒力基因的遗传演化与致病性分析

畜牧兽医学报 2023,54(8):3435-3444A c t a V e t e r i n a r i a e t Z o o t e c h n i c a S i n i c ad o i :10.11843/j.i s s n .0366-6964.2023.08.028开放科学(资源服务)标识码(O S I D ):鸡传染性喉气管炎病毒W F 03株关键抗原基因和毒力基因的遗传演化与致病性分析张翼1,2,王晨燕1,杨骁2,邵国青1,3,侯博1*(1.福建省农业科学院畜牧兽医研究所/福建省畜禽疫病疾病防治工程技术研究中心,福州350013;2.默沙东(宁波)动物保健科技有限公司,宁波315336;3.江苏省农业科学院兽医研究所,南京210014)摘要:本研究旨在分析近年分离自鸡传染性喉气管炎病毒(I L T V )活疫苗免疫失败鸡群的I L T V W F 03株的部分关键抗原基因及毒力基因的变异情况和对鸡的致病力,为I L T V 活疫苗的有效性评价提供一个稳定㊁可靠的攻毒毒株㊂通过I L T V 临床分离株W F 03的部分免疫原性基因和毒力相关基因进行克隆和测序,并与疫苗株和早期分离株进行遗传进化分析,同时在28和56日龄S P F 鸡测定了该毒株的致病力㊂遗传进化分析结果表明:W F 03株主要免疫原性基因和毒力相关基因与我国早期的分离株和现在部分商品化的疫苗株处在同一个大的进化分支,亲缘关系较近㊂致病力试验显示W F 03株以104.5㊁104.0㊁103.5和103.0E I D 50㊃羽-1的剂量感染28日龄S P F 鸡后发病率分别为100%㊁100%㊁95%和85%,死亡率分别为45%㊁30%㊁35%和10%;以104.0E I D 50㊃羽-1的剂量感染56日龄S P F 鸡的发病率和死亡率分别为100%和27.3%;所有发病鸡临床症状主要表现为咳血㊁呼吸困难㊁张口呼吸等典型的喉气管炎发病症状㊂结果表明:I L T V 临床分离株W F 03主要抗原基因和毒力基因与我国早期分离株和疫苗株相比未发生明显变异,属于强致病力毒株,其对S P F 鸡的发病能力稳定,在104.5㊁104.0㊁103.5和103.0E I D 50㊃羽-1攻毒剂量时均可以使80%以上的鸡发病㊂结果提示目前我国鸡群中存在着强致病力I L T V 毒株流行,且W F 03株可以作为疫苗效力评价的攻毒毒株,这将丰富我国当前I L T V 毒株的流行病学数据库㊂关键词:传染性喉气管炎病毒;遗传特征;致病力中图分类号:S 852.659.1 文献标志码:A 文章编号:0366-6964(2023)08-3435-10收稿日期:2022-11-04基金项目:福建省农业科学院科技创新团队(C X T D 2021014-3);福建省 5511 协同创新工程(X T C X G C 2021008)作者简介:张翼(1984-),男,黑龙江哈尔滨人,硕士,兽医师,主要从事兽医传染病防控技术研究,E -m a i l :z h a n g y i 516@h o t m a i l .c o m *通信作者:侯博,主要从事兽医传染病防控技术研究,E -m a i l :h o u b o h o u w e i @126.c o mP h y l o g e n e t i c A n a l y s i s a n d P a t h o g e n i c i t y of a n A v i a n I n f e c t i o u s L a r y n go t r a c h e i t i s V i r u s S t r a i n W F 03Z H A N G Y i 1,2,WA N G C h e n y a n 1,Y A N G X i a o 2,S H A O G u o q i n g 1,3,HO U Bo 1*(1.F u j i a n A n i m a l D i s e a s e C o n t r o l T e c h n o l o g y D e v e l o p m e n t C e n t e r ,I n s t i t u t e o f A n i m a l H u s b a n d r y a n d V e t e r i n a r y M e d i c i n e ,F u j i a n A c a d e m y o f A g r i c u l t u r a l S c i e n c e s ,F u z h o u 350013,C h i n a ;2.M S D (N i n g b o )A n i m a l H e a l t h T e c h n o l o g y C o .,L t d .,N i n g b o 315336,C h i n a ;3.I n s t i t u t e o f V e t e r i n a r yM e d i c i n e ,J i a n g s u A c a d e m y o f A g r i c u l t u r a l S c i e n c e s ,N a n j i n g 210014,C h i n a )A b s t r a c t :T h i s e x p e r i m e n t a n a l y z e d t h e v a r i a t i o n s o f k e y a n t i g e n g e n e s a n d v i r u l e n c e g e n e s o f i n f e c t i o u s l a r y n go t r a c h e i t i s v i r u s (I L T V )W F 03s t r a i n i s o l a t e d f r o m c h i c k e n s t h a t f a i l e d t o b e i mm u n i z e d w i t h l i v e v a c c i n e .A n d a l s o ,t h e v i r u l e n c e o f W F 03w a s t e s t e d i n S P F c h i c k e n s .T h ep h y l o g e n e t i c t r e e o f k e y i mm u n o ge n i c g e n e s a n d v i r u l e n c e r e l a t e d g e n e s of c l i n i c a l I L T V i s o l a t e W F 03w a s p e r f o r m e d ,a n d t h e p a t h og e n i c i t y o f thi s s t r a i n i n f e c t e d 28a n d 56d a ys -o l d S P F畜牧兽医学报54卷c h i c k e n s w a s t e s t e d.T h e r e s u l t s s h o w e d t h a t t h e k e y i mm u n o g e n i c g e n e s a n d v i r u l e n c e r e l a t e d g e n e s o f t h e W F03s t r a i n w e r e i n t h e s i m i l a r i t y e v o l u t i o n a r y b r a n c h w i t h t h e e a r l y i s o l a t e s f r o m C h i n a a n d s o m e c o mm e r c i a l i z e d v a c c i n e s t r a i n s.T h e p a t h o g e n i c i t y e x p e r i m e n t s h o w e d t h a t w h e n t h e28d a y s-o l d S P F c h i c k e n w e r e c h a l l e n g e d w i t h W F03s t r a i n w i t h t h e d o s e s o f104.5,104.0, 103.5a n d103.0E I D50,r e s p e c t i v e l y,t h e m o r b i d i t y w a s100%,100%,95%a n d85%, r e s p e c t i v e l y,a n d t h e m o r t a l i t y w a s45%,30%,35%a n d10%,r e s p e c t i v e l y.T h e m o r b i d i t y a n d m o r t a l i t y t o56d a y s-o l d S P F c h i c k e n s c h a l l e n g e d w i t h104.0E I D50d o s e w e r e100%a n d27.3%, r e s p e c t i v e l y.T h e m a i n c l i n i c a l s y m p t o m s w e r e h e m o p t y s i s,d y s p n e a,m o u t h b r e a t h i n g w i t h s a m e a s t y p i c a l s y m p t o m s o f l a r y n g o t r a c h e i t i s.T h e r e s u l t s s h o w e d t h a t t h e c l i n i c a l s t r a i n W F03 o f I L T V w a s a h i g h v i r u l e n t s t r a i n a n d c a n b e u s e d a s a c h a l l e n g e s t r a i n f o r v a c c i n e e f f i c a c y,a n d i t s g e n e t i c c h a r a c t e r i s t i c s w e r e n o t s i g n i f i c a n t l y d i f f e r e n t f r o m t h o s e o f e a r l y i s o l a t e s a n d v a c c i n e s t r a i n s i n C h i n a.T h e r e s u l t s s u g g e s t e d t h a t h i g h v i r u l e n t s t r a i n s w e r e p r e v a l e n t i n C h i n e s e f l o c k s,w h i c h w i l l e n r i c h t h e e p i d e m i o l o g i c a l d a t a b a s e o f c u r r e n t I L T V s t r a i n s i n C h i n a.K e y w o r d s:i n f e c t i o u s l a r y n g o t r a c h e i t i s v i r u s;g e n e t i c s;p a t h o g e n i c i t y*C o r r e s p o n d i n g a u t h o r:HO U B o,E-m a i l:h o u b o h o u w e i@126.c o m传染性喉气管炎病毒(i n f e c t i o u s l a r y n g o t r a-c h e i t i s v i r u s,I L T V)属于疱疹病毒科㊁α疱疹病毒亚科传染性喉气管炎病毒属㊂I L T V感染鸡后引起鸡喉气管炎(i n f e c t i o u s l a r y n g o t r a c h e i t i s,I L T),主要危害成年鸡和产蛋鸡,造成死亡和/或产蛋下降,种鸡的I L T发病呈世界性分布,世界上许多国家都有该病的发病报道,该病在我国近几年也呈散发流行趋势,不断有养殖场发生I L T V感染的报道[1-4]㊂临床上严重的I L T V感染往往表现为呼吸困难㊁喘气㊁咳血等,具有很高的致死性,而温和型感染主要表现为黏液性气管炎㊁鼻炎㊁结膜炎㊁不愿活动和低死亡率等多种形式,不论是严重感染还是温和型感染均给生产带来了严重的损失和威胁㊂在集约化养禽的发达国家,已使用弱毒疫苗控制了蛋鸡的I L T,但有报道称在2019年4月以来我国山东省多个地区发生了多起商品肉鸡㊁地方品种肉鸡等感染I L-T V的病例,临床主要表现为严重的咳血㊁呼吸困难以及呼吸道粘膜出血和充血[3],表明I L T V感染肉鸡目前在我国呈现出散发和多发现象㊂I L T V的基因组大小约为155k b,是一种双链D N A病毒,与其他疱疹病毒一样,基因组包括长独特区㊁短独特区以及两端的反向重复序列等结构,编码大量的糖蛋白以及与毒力和病毒复制有关的蛋白㊂g B和U L32基因编码病毒粒子的表面糖蛋白,其中g B不仅是病毒感染所必需的糖蛋白,而且是一种主要的保护性抗原,可诱导体液免疫[5],并且在病毒感染过程中诱导膜融合和病毒穿入[6-7]㊂糖蛋白g D作为疱疹病毒的受体对于病毒结合到敏感细胞的表面是必需的[8-9]㊂糖蛋白g B和g D也是I L-T V等疱疹病毒的保护性抗原,可诱导中和抗体和细胞免疫反应,是重要的免疫原蛋白[10-11]㊂而糖蛋白g E/g I对于I L T V在细胞与细胞之间扩散具有显著的促进作用,缺失g E/g I的I L T V病毒噬斑形成能力显著下降,仅能够在单细胞内增殖[12]㊂T K基因是I L T V的主要毒力基因[13],缺失T K可以使I L T V的致病力减弱但仍然保持良好的免疫原性[14]㊂I C P4基因参与激活病毒早/晚期基因的表达,与病毒毒力存在关联[15]㊂糖蛋白g C是I L T V 主要的表面抗原蛋白,对病毒的复制是非必需的,但缺失g C可以使I L T V对鸡的致病力减弱,并且增强先天性和特异性免疫反应[16]㊂糖蛋白g G是I L-T V的毒力因子,缺失g G可以使病毒毒力显著减弱[17]㊂此外还有报道糖蛋白g D㊁g B㊁g H㊁g L㊁g K㊁g M㊁g N等参与了疱疹病毒致病性的形成[18-21]㊂I L T V野毒株的毒力差异很大,部分毒株感染鸡后具有很高的发病率和致死率,属于强致病力毒株,而感染后症状轻微或不明显㊁无致死率的毒株属于温和型毒株,因此及时了解我国当前散发的I L-T V毒株的致病性和主要抗原基因和毒力基因的系统发育和进化关系,以及筛选一株攻毒后发病稳定的强毒株用于I L T V活疫苗的效力检验,对于做好I L T的防控具有重要的临床意义㊂本研究对1株临床I L T V分离株的主要免疫原性基因和毒力基因进行遗传进化分析,了解其与疫苗株之间的进化亲缘63438期张翼等:鸡传染性喉气管炎病毒W F 03株关键抗原基因和毒力基因的遗传演化与致病性分析关系,并在28日龄和56日龄S P F 鸡测定了该毒株的致病力,其研究结果为我国有效预防和控制I L -T V 的感染提供了临床参考㊂1 材料与方法1.1 主要试剂与材料含1%庆大霉素的胰蛋白磷酸盐肉汤(T P B ),购自S i gm a 公司;1-5T M2ˑH i g h F i d e l i t y ma s t e r M i x 购自M C L A B 公司;D N A 胶回收试剂盒购自T h e r m o F i s h e r 公司,平末端载体p C l o n e 007试剂盒购自擎科生物科技有限公司;D N A 提取试剂盒购自Q i a ge n 公司;舒泰购自法国维克有限公司;S P F 鸡胚购自济南斯帕法斯家禽有限公司;56和28日龄S P F 鸡购自济南赛斯家禽科技有限公司㊂P C R 引物由生工生物工程(上海)股份有限公司合成(表1)㊂表1 P C R 扩增使用的引物名称和序列T a b l e 1 P r i m e r s n a m e a n d s e q u e n c e o f t h i s s t u d y us e d 基因名称G e n e n a m e引物序列(5'ң3')P r i m e s e qu e n c e 产物长度/b pS i z e g K F :G C A T A C A C A A C G G T T G A G A C ;R :G G A A T C T A C T C G C C A G G A C G 2216g G F :C T G A C C G C T A G C C C A T A C A G ;R :T C C A G T G C A A G T T C G T C C T C1416g E F :C A G T A T C G G A C A C G G A C T G G ;R :C G G G T T G A C T G G A A A C G G T A1894g C F :A G A C T G T C G C C T C G A A A C T G ;R :T T G G T C A A C A C T G C A A A G C G1983gL F :A C T G T A A C A G C G T C T C A C C G ;R :T A C C G G A G A C A G A A G A G G G G1377g N F :A T T G C A A G G A A T C G A G G C C A ;R :T A C T T C T T C G C C T C C T C C G A692g M F :C C C C A C A A C C A T C G A C A G A A ;R :G C T G A G A T C G C A T C C G T A C A 1722g D F :T A C C G C T G G C C A A A G T A C A G ;R :T A G T C T G T T C G A G G C G G A G A1850g B F :G A C A G T C G A T G C T T C T A T G C C ;R :A T G C A A T C C T A C A T C G C C G T [22]2799g H F :T G C T T C T T T G A C G G A G C C A T ;R :C C G A G A C G G C A A G T T C T C T T2617g I F :C C T C C A G A A G A C T C G G A G G A ;R :C C G G A A C G T T T C T G G G A T C A1789T K F :C A T G A C T A C A A T T T T G C C C ;R :A C G A A C C C A T A T C A G C A T T[22]1251U L 32F :G G T A T G A G A T G T A G C G A T G G ;R :G G T A G T T T A G T T T A G C A T C G G T A[22]2023I C P 41F :T G C T T T T C C T G C A G C G T C T A ;R :T C T T G T C T C A G A A C G T G C C C2244I C P 42F :C G T G A G T G A G G A A G G G G ;R :G G T A C A G A C G G G T C G G T [22]2668I C P 43F :G G C A T C C C G T A C T C T T C G A G ;R :T C A C T C G G G T C T T G T T C T G C12721.2 病毒的繁殖与E I D 50测定I L T V 临床分离株W F 03株由福建省农业科学院畜牧兽医研究所保存㊂该病毒株分离自免疫了I L T V 细胞源弱毒活疫苗的黄羽肉鸡群,鸡群在40~50日龄出现了以咳血和死亡为特征的疑似I L T ,发病率较高,部分鸡出现死亡㊂采用有限稀释法纯化病毒,将保存的I L T V 病毒液以T P B 连续10倍稀释,取10-1㊁10-2㊁10-3㊁10-4㊁10-5㊁10-6稀释度采用鸡胚尿囊膜(C A M )方式接种10日龄S P F鸡胚,每个稀释度接种6枚,接种量为0.2m L ㊃胚-1,弃去24h 内死亡鸡胚,继续孵化7d ,每日检查鸡胚存活情况,7d 后对存活鸡胚检查尿囊膜,死亡鸡胚和尿囊膜出现灰白色斑的鸡胚判定为阳性㊂取最高稀释度的阳性尿囊膜以1ʒ5的比例加入预冷的P B S 进行研磨,反复冻融3次,12000r ㊃m i n -1,4ħ离心10m i n,取上清病毒液采用同样方式继续纯化2次㊂将纯化第3次收获的阳性病毒液命名为W F 03株P 0代㊂采用C AM 方式接种10日龄S P F 鸡胚7d后收获尿囊膜并按照上述方法制备病毒液,记为P 1代;依次传代制备不同代次的病毒液,分别记为P 2代㊁P 3代等㊂将待测E I D 50的病毒液以TP B 连续稀释成10-3.6㊁10-4.2㊁10-4.8㊁10-5.4和10-6.05个稀释度,从高稀释度到低稀释度经C AM 接种病毒液,每个稀释度接种6枚胚,每胚接种0.2m L ,置37ħ继续孵化7d ,每天检查鸡胚存活情况并作记录㊂同时取6个S P F 鸡胚以同样方式接种T P B 作为阴性对照㊂弃去接种后24h 内的死亡鸡胚,每个稀释度应至少有4个鸡胚存活,阴性对照鸡胚在接种后第2~7天不应该有死亡出现,试验才有效㊂接种24h 后死亡的鸡胚判定感染阳性,接种第7天后对所有存活鸡胚进行检查,当鸡胚尿囊膜有明显增厚出现白色病7343畜牧兽医学报54卷斑,判为感染阳性㊂采用R e e d-M u e n c h法计算鸡胚半数感染量E I D50,重复测定3次,至少2次测定试验成立,结果才有效㊂1.3I L T V主要基因的扩增与测序将300m L I L T V P5代病毒液以45000r㊃m i n-1, 4ħ离心4.5h,弃去上清后将沉淀物使用D N A提取试剂盒按照说明书提取基因组D N A后,采用表1中的引物分别对免疫原性基因g B㊁U L32㊁g D㊁g C 和参与毒力形成的基因T K㊁I C P4㊁g G㊁g E㊁g I㊁g H㊁g L㊁g K㊁g M㊁g N进行P C R扩增㊂反应体系: 1-5T M2ˑH i g h F i d e l i t y m a s t e r M i x25μL,上㊁下游引物各1μL,D N A模板2μL,d d H2O21μL;反应条件:98ħ2m i n;98ħ10s,56ħ15s,72ħ30s,共30个循环;72ħ5m i n㊂扩增结束后,将P C R产物进行1%琼脂糖凝胶电泳,将目的条带切胶并利用D N A胶回收试剂盒回收P C R产物㊂将回收到的P C R产物连接到平末端载体p C l o n e007中转化感受态细胞D H5α后涂布含氨苄抗生素的平板,每个基因均筛选5个克隆子(菌液)送生工生物工程(上海)股份有限公司进行测序㊂1.4关键抗原基因和毒力基因遗传进化分析从N C B I下载I L T V国内外主要分离株和疫苗株的基因组序列(表2),使用D N A S t a r软件和G e-n e i o u s软件对参考毒株和分离毒株的免疫原性基因g B㊁U L32㊁g D㊁g C和毒力相关基因T K㊁I C P4㊁g G㊁g E㊁g I㊁g H㊁g L㊁g K㊁g M㊁g N进行分析和串表2I L T V参考毒株信息T a b l e2I L T V r e f e r e n c e s t r a i n i n f o r m a t i o n菌株名S t r a i n n a m eG e n B a n k登陆号G e n B a n k a c c e s s i o n N o.国家C o u n t r y年Y e a rV F A R-043MG775218秘鲁P e r u2014 U S D A r e f e r e n c e J N542534美国U S A2012 L J S09J X458822中国C h i n a2009 WG J X458823中国C h i n a2012 C S W-1J X646899澳大利亚A u s t r a l i a1970 30678/14/K o MH937565韩国S o u t h K o r e a2014 A C C78/J N804826澳大利亚A u s t r a l i a2011 4787/80K P677885意大利I t a l y1980 193435/07K P677883意大利I t a l y2007 A489K Y423284美国U S A2017 27_C S W-1/V1-99M F156852澳大利亚A u s t r a l i a2016 J2M F417808美国U S A2017 R u s/C k/P e n z a/2013/2701M F405080俄罗斯R u s s i a2013 I L T V.319/19MK895002澳大利亚A u s t r a l i a2019 c k/C H/L H L J/120305M T876619中国C h i n a2012 40798/10/K o MH937566韩国S o u t h K o r e a2010 7b MN335811澳大利亚A u s t r a l i a2016 C A N/Q C-2236832O L354140加拿大C a n a d a2019 C A N/Q C-2301711O K646550加拿大C a n a d a2020 C A N/O N-2462242O K624781加拿大C a n a d a1987 C A N/O N-2462241O K573459加拿大C a n a d a2021 L i v e a t t e n u a t e d s t r a i nA20J N596963澳大利亚A u s t r a l i a2011 K317J X458824中国C h i n a2012 L T B l e n J Q083493美国U S A2011 L a r y n g o V a c J Q083494美国U S A2011 P o u l v a c I L T(R)K P677882意大利I t a l y2015 R u s/C k/T a t a r s t a n/2009/1643M F405079俄罗斯R u s s i a2017 N o b i l i s L a r i n g o v a c(R)K P677881意大利I t a l y2015 83438期张翼等:鸡传染性喉气管炎病毒W F03株关键抗原基因和毒力基因的遗传演化与致病性分析联拼接,对拼接后的序列采用M E G A7软件中的C l a s t a l W算法对所有序列进行多重序列比对后,以邻接(n e i g h b o r-J o i n i n g,N J)法中重复数1000计算b o o t s t r a p值构建免疫原性基因g B-g C-g D-U L32和毒力相关基因g E-g I-T K-I C P4-g G-g H-g K-g L-g M-g N的进化树㊂1.5病毒对S P F鸡的致病力分析将测定过E I D50的W F03株P5代病毒液用T B P稀释后以气管攻毒途径对S P F鸡进行感染㊂选取90羽28日龄S P F鸡随机分为5组,其中第1~4组(每组20羽)以气管内注射途径分别以104.5㊁104.0㊁103.5和103.0E I D50㊃羽-1的剂量接种W F03株0.2m L;第5组(10羽)经气管内注射途径接种0.2m L的T P B稀释液,作为阴性对照㊂另取8周龄S P F鸡,共分为2组,每组11只鸡,其中1组用W F03株以104.0E I D50㊃只-1的剂量进行气管内注射0.2m L,另一组以T B P稀释液为空白对照组㊂攻毒后,每日对所有鸡进行观察两次,共观察14d,每天记录每羽鸡的发病情况,包括死亡㊁咳血㊁呼吸症状㊁眼部症状和精神状态等㊂死亡鸡随时解剖,未死亡鸡在试验结束后采用舒泰过量麻醉致死后统一剖检,重点观察喉头㊁气管等病理变化,包括气管内有无血块㊁黄色酪状物栓塞㊂2结果2.1I L T V繁殖特性I L T V W F03株通过C AM途径采用有限稀释法接种10日龄S P F鸡胚,在接种鸡胚7d后,幸存胚的尿囊膜出现大量灰白色病灶或尿囊膜出现大量的灰白色痘斑(图1)㊂测定纯化后病毒的E I D50,其P1㊁P2㊁P3和P5代的病毒含量分别为105.4㊁105.8㊁105.5和105.89E I D50㊃m L-1㊂2.2I L T V关键抗原基因和毒力基因的遗传进化分析对串联的免疫原性基因g B-g C-g D-U L32的遗传进化分析显示:W F03株与中国2009年的分离株L J S09株㊁2012年的分离株c k/C H/L H L J/ 120305株㊁疫苗株K317和N o b i l i s L a r i n g o v a c(R)处在一个大的进化分支上,并且与疫苗株P o u l v a c I L T(R)㊁L a r y n g o V a c和L T B l e n处在同一个分支上且亲缘关系最近(图2)㊂通过对W F03株与参考毒株的基因序列比对显示:g B基因与参考毒株的相似性为99.67%~99.93%,g C基因与参考毒株的相似性为99.75%~100.00%,g D基因与参考毒株的相似性为99.14%~100.00%,U L32基因与参考毒株的相似性为99.90%~100.00%㊂I L T V接种鸡胚7d后尿囊膜显示灰白色痘斑(右),左侧为正常尿囊膜对照G r a y w h i t e a c n e-s p o t(r i g h t)o f a l l a n t o i c m e m b r a n e7d a y s a f t e r i n o c u l a t i o n,t h e l e f t i s n o r m a l a l l a n t o i c m e m b r a n e c o n t r o l图1I L T V W F03株接种鸡胚7d后尿囊膜病理变化F i g.1T h e p a t h o l o g i c a l c h a n g e s o f a l l a n t o i c m e m b r a n e7d a y sa f t e r i n o c u l a t i o n o f c h i c k e n e mb r y o w i t h I L T V W F03s t r a i n对串联的毒力相关基因g E-g I-T K-I C P4-g G-g H-g K-g L-g M-g N的遗传进化分析显示:W F03株与野毒株进化关系表现为与韩国2010年分离的40798/10/K o毒株亲缘关系最近,其次与我国2012年分离株c k/C H/L H L J/120305㊁2009年分离株L J S09或澳大利亚2016年分离株7b株的亲缘关系较近;W F03株与疫苗株的进化关系表现为与L T B l e n株的亲缘关系最近,与K317和N o b i l i s L a r i n-g o v a c(R)的亲缘关系次之(图3)㊂通过W F03株与参考毒株的基因序列比对显示:g E基因与参考毒株的相似性为99.47%~100.00%,g I基因与参考毒株的相似性为99.05%~100.00%,T K基因与参考毒株的相似性为99.68%~100.00%,I C P4基因与参考毒株的相似性为98.71%~99.96%,g G 基因与参考毒株的相似性为98.73%~100.00%, g H基因与参考毒株的相似性为99.58%~ 100.00%,g K基因与参考毒株的相似性为99.59%~100.00%,g L基因与参考毒株的相似性为99.84%~100.00%,g M基因与参考毒株的相似性为99.77%~100.00%,g N基因与参考毒株的相似性为99.28%~100.00%㊂2.3I L T V对S F P鸡的致病力W F03株以104.5㊁104.0㊁103.5和103.0E I D50㊃羽-1的剂量感染28日龄S P F鸡后发病率分别为100% (20/20)㊁100%(20/20)㊁95%(19/20)和85%(17/9343畜牧兽医学报54卷ә.疫苗株;һ.本研究分离株ә.V a c c i n e s t r a i n s ;һ.I s o l a t e i n o u r s t u d y图2 免疫原性基因的遗传进化分析F i g .2 P h y l o g e n e t i c t r e e a n a l y s i s o f g e n e s r e l a t e d t o i m m u n o g e n i c i t y20),死亡率分别为45%(9/20)㊁30%(6/20)㊁35%(7/20)和10%(2/20),阴性对照组未发生特异性临床症状㊂在W F 03株攻毒后2d 开始陆续出现临床症状,持续期均超过2d 并且食欲废绝,相继出现精神沉郁㊁咳血㊁呼吸困难㊁张口呼吸等典型的I L T 发病症状㊂对死亡鸡解剖后发现气管中有大量血液或血凝块㊂其余存活鸡从攻毒后第7天开始临床症状逐步消失㊂结果表明W F 03株可以导致28日龄S P F 鸡出现较高的发病率和死亡率㊂W F 03株感染56日龄S P F 鸡后2d 开始陆续出现临床症状,持续期均超过3d 并且食欲废绝,第5天时死亡3只,以中度至重度气喘(11/11)(图4)㊁咳血(7/11)(图4)㊁中度至重度呼吸困难(7/11)为主要临床症状㊂对第5天死亡鸡解剖发现气管中有大量血液或血凝块(图5),其余存活鸡到攻毒后9日临床症状消失㊂结果表明W F 03株可以导致56日龄S P F 鸡100%(11/11)的发病率和27.3%(3/11)的死亡率㊂因此结果表明W F 03株对于S P F 鸡的具有较强的致病力,其以不同剂量感染均可以使80%的鸡发病㊂3 讨论在我国,鸡胚源(c h i c k e n e m b r y o o r i gi n ,C E O )和组织细胞源(t i s s u e c u l t u r e o r i gi n ,T C O )2种弱毒活疫苗已批准用于I L T 预防和控制,但在近几年仍有养殖场发生I L T V 感染的报道,尤其在肉鸡中也发现了I L T V 感染的病例[1-4]㊂糖蛋白g B ㊁gD 和g C 以及U L 32蛋白是I L T V 重要的保护性抗原,可以诱导中和抗体和细胞免疫反应,并且还可以增强先天性和特异性免疫反应[5,10-11,16],此外糖蛋白g B 和g D 还参与病毒黏附㊁膜融合和穿入的过程[6-9],而糖蛋白g C 虽不是病毒复制必需基因,但04438期张翼等:鸡传染性喉气管炎病毒W F 03株关键抗原基因和毒力基因的遗传演化与致病性分析ә.疫苗株;һ.本研究分离株ә.V a c c i n e s t r a i n s ;һ.I s o l a t e i n o u r s t u d y图3 毒力相关基因的遗传进化分析F i g .3 P h y l o g e n e t i c t r e e a n a l ys i s o f g e n e s r e l a t e d t o v i r u l e n c e (B)图4 攻毒后出现张口呼吸和咳血(左)和未感染对照鸡(右)F i g .4 O p e n m o u t h b r e a t h i n g a n d h e m o p t y s i s a f t e r c h a l l e n g e (l e f t )a n d c o n t r o l w i t h o u t c h a l l e n g e (r i gh t )缺失g C 可以使I L T V 对鸡的致病力减弱[16],因此g B ㊁gC 和gD 在I L T V 致病力的形成中同样发挥着重要作用㊂本研究通过对I L T V W F 03分离株的免疫原性基因g B -g C -gD -U L 32的遗传进化分析发现W F 03分离株与我国早期I L T V 分离株的亲缘关系较近,其主要抗原基因与现有主要疫苗毒株K 317㊁P o u l v a c I L T (R )㊁L a r y n go V a c 和L T B l e n 的亲缘关系均较近,提示现有疫苗能够对目前的I L T V 分离株提供有效保护,然而在免疫鸡群中仍然具有I L T 的散发病例㊂目前预防I L T V 感染的疫苗不仅有C E O 和T C O 两种弱毒活疫苗,基因工程载体疫苗也已经商品化,由于不同疫苗制备工艺不同,且I L T V 临床分离株的毒力强弱差异较大,因此选用何种疫苗免疫,应该在使用前进行有效性评估㊂在疱疹病毒(包括I L T V )中,糖蛋白g E ㊁gI 和1443畜牧兽医学报54卷图5 攻毒后死亡鸡剖检气管可见大量新鲜血液或血凝块(左)和未感染对照鸡气管(右)F i g .5 A l a r g e a m o u n t o f f r e s h b l o o d o r b l o o d c l o t s w a s f o u n d i n t r a c h e a l n e c r o p s y o f d e a d c h i c k e n s a f t e r c h a l l e n ge a n d t r a c h e a w i t h o u t c h a l l e n g e (r i gh t )gG 以及T K 基因缺失可以使病毒毒力显著减弱,并且使病毒保持良好的免疫原性[12-14,17]㊂I C P 4基因参与激活病毒早/晚期基因的表达,与病毒毒力存在关联[15]㊂此外还有报道糖蛋白g H ㊁g L ㊁gK ㊁g M ㊁gN 等也参与了疱疹病毒致病性的形成[18-21]㊂而在本研究中通过28和56日龄S P F 鸡的感染试验结果表明以104.0E I D 50的剂量攻毒后表现出严重的咳血和死亡,W F 03株属于强致病力毒株,但通过对毒力相关基因g E -g I -T K -I C P 4-g G -g H -gK -g L -g M -gN 的进化树分析,其亲缘关系与韩国2010年分离的40798/10/K o 毒株亲缘关系最近,其次分别与我国2012年分离株c k /C H /L H L J/120305㊁2009年分离株L J S 09和澳大利亚2016年分离株7b 株的亲缘关系较近,并且与疫苗株中的L T B l e n 和K 317株的亲缘关系同样较近㊂有报道根据T K ㊁I C P 4㊁g B 和U L 32基因核苷酸序列分析结果,近年新分离的I L T V B T 株与我国早期L J S 09毒株等属于同一进化分支,未出现较大变异,但以104.0E I D 50的剂量气管内攻毒2周龄S PF 鸡未出现死亡,仅有少部分鸡出现结膜分泌物,剖检显示喉头气管正常[22],但早期毒株L J S 09株对4周龄鸡以102E I D 50攻毒,急性症状出现在感染后4~6d ,表现为呼吸困难,并且发病率和死亡率分别为80%和10%[23],因此本试验I L T V W F 03株感染28日龄和56日龄S P F 鸡后的发病时间㊁比例㊁症状与其他I L T V 强致病力毒株感染情况基本一致㊂因此本研究通过对I L T V 主要基因的遗传特征分析表明目前I L T V 临床分离株与我国早期的分离株在毒力基因的遗传关系上无显著的差异,I L T V 未发生较大的遗传变异现象,但分离株的致病力具有较大差异,在临床上可能呈现出温和型和急性型I L T V 共存的局面,目前现有疫苗可能仍对当前I L T V 强致病力毒株的流行具有保护作用㊂此外,目前由于I L T V 疫苗免疫效力评估尚没有标准株使用,不同生物制品厂使用不同I L T V 强毒株对疫苗进行效力检验㊂然而,I L T V 野毒株的毒力差异很大,有的毒株具有很高的发病率和致死率属于强致病力毒株,而有的毒株感染后仅表现出结膜炎等轻微症状或症状不明显,对于筛选一株遗传稳定㊁发病可靠㊁临床症状易于观察的强毒株进行I L T V 疫苗的效力检验是非常有必要的,也是非常重要的㊂W F 03株以104.5㊁104.0㊁103.5和103.0E I D 50㊃羽-1的剂量感染28日龄S P F 鸡后至少有80%的鸡发病,且以咳血㊁死亡为主要临床症状,对56日龄S P F 鸡以104.0E I D 50㊃羽-1的剂量攻毒后,100%的鸡发病,主要以气喘㊁呼吸困难和咳血为主要临床症状㊂因此W F 03株可作为疫苗效力检验用强毒株,其发病稳定,临床症状易于观察㊂4 结论本研究通过28和56日龄S P F 鸡的人工感染试验结果表明I L T V W F 03分离株具有很强的致病力,对28和56日龄鸡的致死性也较强,但通过免疫原性基因g B -g C -gD -U L 32和毒力相关基因gE -g I -T K -I C P 4-g G -g H -g 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e n e a n d g B g e n e of i n f e c t i o u sl a r y n g o t r a c h e i t i s v i r u s[J].C h i n e s e J o u r n a l o fV e t e r i n a r y M e d i c i n e,2015,51(2):6-8,12.(i nC h i n e s e)[6] C O N N O L L Y S A,J A C K S O N J O,J A R D E T Z K Y TS,e t a l.F u s i n g s t r u c t u r e a n d f u n c t i o n:a s t r u c t u r a lv i e w o f t h e h e r p e s v i r u s e n t r y m a c h i n e r y[J].N a t R e vM i c r o b i o l,2011,9(5):369-381.[7] S P E A R P G,L O N G N E C K E R R.H e r p e s v i r u s e n t r y:a nu p d a t e[J].J V i r o l,2003,77(19):10179-10185. [8] L U G W,Z H A N G N,Q I J X,e t a l.C r y s t a l s t r u c t u r eo f h e r p e s s i m p l e x v i r u s2g D b o u n d t o n e c t i n-1r e v e a l s ac o n s e r v ed m o de of r e c e p t o r r e c og n i t i o n[J].J V i r o l,2014,88(23):13678-13688.[9] D I G I O V I N E P,S E T T E M B R E E C,B HA R G A V AA K,e t a l.S t r u c t u r e o f h e r p e s s i m p l e x v i r u sg l y c o p r o t e i n D b o u n d t o t h e h u m a n r e c e p t o r n e c t i n-1[J].P L o S P a t h o g,2011,7(9):e1002277. [10] L A Z E A R E,WH I T B E C K J C,P O N C E-D E-L E O NM,e t a l.A n t i b o d y-i n d u c e d c o n f o r m a t i o n a l c h a n g e si n h e r p e s s i m p l e x v i r u s g l y c o p r o t e i n g D r e v e a l n e wt a r g e t s f o r v i r u s n e u t r a l i z a t i o n[J].J V i r o l,2012,86(3):1563-1576.[11] Z HA O W,S P A T Z S,Z HA N G Z Y,e t a l.N e w c a s t l e d i s e a s e v i r u s(N D V)r e c o m b i n a n t se x p r e s s i n g i nf e c t i o u s l a r y ng o t r a ch ei t i s v i r u s(I L T V)g l y c o p r o t e i n s g B a n d g D p r o t e c t c h i c k e n s a g a i n s tI L T V a n d N D V c h a l l e n g e s[J].J V i r o l,2014,88(15):8397-8406.[12] D E V L I N J M,B R OWN I N G G F,G I L K E R S O N JR.A g l y c o p r o t e i n I-a n d g l y c o p r o t e i n E-d e f i c i e n tm u t a n t o f i n f e c t i o u s l a r y n g o t r a c h e i t i s v i r u s e x h i b i t si m p a i r e d c e l l-t o-c e l l s p r e a d i n c u l t u r e d c e l l s[J].A r c hV i r o l,2006,151(7):1281-1289.[13] HA N M G,KW E O N C H,MO I P,e t a l.P a t h o g e n i c i t y a n d v a c c i n e e f f i c a c y o f a t h y m i d i n ek i n a s e g e n e d e l e t e d i n f e c t i o u s l a r y n g o t r a c h e i t i s v i r u se x p r e s s i n g t h e g r e e nf l u o r e s c e n t p r o t e i ng e n e[J].A r c h V i r o l,2002,147(5):1017-1031.[14] S C HN I T Z L E I N W M,W I N A N S R,E L L S WO R T HS,e t a l.G e n e r a t i o n o f t h y m i d i n e k i n a s e-d e f i c i e n tm u t a n t s o f i n f e c t i o u s l a r y n g o t r a c h e i t i s v i r u s[J].V i r o l o g y,1995,209(2):304-314.[15]J OHN S O N M A,T Y A C K S G,P R I D E A U X C,e ta l.N u c l e o t i d e s e q u e n c e o f i n f e c t i o u s l a r y n g o t r a c h e i t i sv i r u s(g a l l i d h e r p e s v i r u s1)I C P4g e n e[J].V i r u sR e s,1995,35(2):193-204.[16] P A V L O V A S P,V E I T S J,B L OHM U,e t a l.I nv i t r o a n d i n v i v o c h a r a c t e r i z a t i o n o f g l y c o p r o t e i n C-d e l e t e d i n f e c t i o u s l a r y n g o t r a c h e i t i s v i r u s[J].J G e nV i r o l,2010,91(4):847-857.[17] D E V L I N J M,B R OWN I N G G F,HA R T L E Y C A,e t a l.G l y c o p r o t e i n G i s a v i r u l e n c ef a c t o r i n i n f e c t i o u sl a r y n g o t r a c h e i t i s v i r u s[J].J G e n V i r o l,2006,87(10):2839-2847.3443畜牧兽医学报54卷[18] V A L L B R A C H T M,R E HWA L D T S,K L U P P B G,e t a l.F u n c t i o n a l r e l e v a n c e of t h e N-t e r m i n a l d o m a i no f p s e u d o r a b i e s v i r u s e n v e l o p e g l y c o p r o t e i n H a n d i t si n t e r a c t i o n w i t h g l y c o p r o t e i n L[J].J V i r o l,2017,91(9):e00061-17.[19] MÖH L B S,C H E N J,P A R K S J,e t a l.E p s t e i n-b a r r v i r u s f u s i o n w i t h e p i t h e l i a lc e l l s t r i g g e r ed b y g Bi s r e s t r i c t e d b y a g L g l y c o s y l a t i o n s i t e[J].J V i r o l,2017,91(23):e01255-17.[20] C HO U L J E N K O D V,J AM B U N A T HA N N,C HO U L J E N K O V N,e t a l.H e r p e s s i m p l e x v i r u s1U L37p r o t e i n t y r o s i n e r e s i d u e s c o n s e r v e d a m o n g a l la l p h a h e r p e s v i r u s e s a r e r e q u i r e d f o r i n t e r a c t i o n s w i t hg l y c o p r o t e i n K,C y t o p l a s m i c v i r i o n e n v e l o p m e n t,a n di n f e c t i o u s v i r u s p r o d u c t i o n[J].J V i r o l,2016,90(22):10351-10361.[21] E L K A S M I I,L I P PÉR.H e r p e s s i m p l e x v i r u s1g Np a r t n e r s w i t h g M t o m o d u l a t e t h e v i r a l f u s i o nm a c h i n e r y[J].J V i r o l,2015,89(4):2313-2323.[22]高昕,罗园园,杨慧明,等.1株鸡传染性喉气管炎病毒的基因组特征与致病性[J].畜牧兽医学报,2022,53(4):1298-1302.G A O X,L U O Y Y,Y A N G H M,e t a l.G e n o m i cc h a r a c t e r i s t i c s a nd p a t h o ge n i c i t y of a n a v i a n i n f e c t i o u sL a r y n g o t r a c h e i t i s v i r u s[J].A c t a V e t e r i n a r i a e tZ o o t e c h n i c a S i n i c a,2022,53(4):1298-1302.(i nC h i n e s e)[23]蒋露,李慧昕,韩宗玺,等.鸡传染性喉气管炎病毒L J S09株致病性的研究[J].中国预防兽医学报,2015,37(3):181-184.J I A N G L,L I H X,HA N Z X,e t a l.P a t h o g e n i c i t ys t u d y o f i n f e c t i o u s l a r y n g o t r a c h e i t i s v i r u s L J S09[J].C h i n e s e J o u r n a l o f P r e v e n t i v e V e t e r i n a r y M e d i c i n e,2015,37(3):181-184.(i n C h i n e s e)(编辑白永平)4443。

Philips Fidelio A5无线Hi-Fi音频扬声器说明书

Philips FidelioA5 wireless Hi-Fi speakerHigh fidelity stereo sound 60W, 4 Hi-Fi driversAnalog-inAW5000All your music in high fidelity sound. Wirelessly. Obsessed with soundGet your music wirelessly, in high fidelity sound with the Philips Fidelio Wireless Hi-Fi speaker. Stream music effortlessly with 4 Hi-Fi drivers to offer a powerful & fully immersive music experience.High fidelity stereo sound•Angled speaker for wider stereo sound•Authentic sound from 4 audiophile-grade drivers•Audio line-inEnjoy your music wirelessly•Enjoy a universe of online music services•Tune into thousands of Internet radio stations•Support music streaming via UPnPIntuitive and easy to use•AirStudio+ Lite to control music from your mobile device•Quick and easy set-upDesign that blends into your interior•Versatile placement to fit in any homeIssue date 2022-06-03Version: 12.11.712 NC: 8670 000 91531EAN: 87 12581 64970 8© 2022 Koninklijke Philips N.V.All Rights reserved.Specifications are subject to change without notice.Trademarks are the property of Koninklijke Philips N.V.or their respective owners. HighlightsA5 wireless Hi-Fi speaker High fidelity stereo sound 60W, 4 Hi-Fi drivers, Analog-inSpecificationsAirStudio+ Lite App Available from the Apple App store and Google Play, the App automatically detects mobile and other compatible devices connected to your home Wi-Fi network, giving you instant control of your music. Control music playback, volume and sound settings, as well as online music services and Internet radio from any device, anywhere.4 audiophile-grade drivers The speaker features two tweeters and two Hi-Fi mid-range drivers. These top-quality components are optimally positioned in order to perfectly deliver dynamically immersive sound.Online music service Getting a subscription to online music services is one of the best ways to get your hands on a variety of music, including the latest releases. You can also access music anytime, anywhere, as long as there is an Internet connection. With the Philips Wireless Hi-Fi, you don't even need to turn on your computer to enjoy the online music service.Internet radio stations The Fidelio Wireless Hi-Fi lets you enjoy more than 30,000 online radio stations from all across the globe. Expand your musical horizons by discovering great stations by genre or by country, or pick your favorite stations from the vast selection available right at your fingertips. It’s super easy too – every Fidelio Wireless Hi-Fi features direct access keys toany station.Sound •Sound Enhancement: FullSound, Treble and Bass Control •Volume Control: up/downLoudspeakers •Loudspeaker types: Integrated •Angled speaker technology •Built-in speakers: 4•Speaker Drivers: 2 x dome tweeters, 2 x 4.25" woofersAudio Playback •Compression format: MP3, WMA, non DRM AAC (m4A), eAAC+, FLAC •Sample frequencies: 8-48 kHz (MP3)•MP3 bit rates: 8-320kpbs and VBR •ID3-tag support •Music Streaming: Internet Radio, Music Services, PC/MAC Connectivity •Wireless connection: Wireless LAN (802.11n), Wireless LAN(802.11g), Wireless LAN(802.11b)•Wireless Universal Plug & Play: UPnP client •LAN wired: Ethernet (RJ 45) 1x •Encryption / security: WEP 128 bit, WEP 64 bit, WPA, WPS-PBC, WPS-PIN, WPA2•Audio Connections: 3.5mm stereo line in -MP3 linkConvenience•Internet Radio preset keys •Wall mountable Smartphone/Tablet App •App name: AirStudio+ Lite, Free download from Apple App store/Google Play Accessories •User Manual: English, French, German, Spanish, Italian, Dutch, Norwegian, Swedish, Finnish, Portuguese, Danish, Russian, Polish (available online)•Warranty Card: Warranty Card •Included accessories: AC Power Cord, Quick start guide, World Wide Warranty leafletDimensions •Product dimensions (WxDxH): 330 x 205 x 258mm •Packaging dimensions (WxDxH): 360 x 249 x 317 mm •Gross Weight: 6.5 lb •Net Weight:5 kg*Napster is available only in Germany and the United Kingdom.。

Fidelity 个性化规划与建议管理账户服务FAQ说明书

Fidelity® Personalized Planning & AdviceFAQWhat is a managed account service?A Fidelity Personalized Planning & Advice managed account develops a risk-managedinvestment strategy that factors in your unique financial situation. We guide youthrough a series of questions to get to know your risk tolerance, financial situation,and how much you may need in retirement. You’ll have access to an exclusivedashboard to track income estimates, your progress toward retirement, and makeupdates based on life events.Is a managed account right for me?A managed account may be just what you're looking for if you:o Want a team of professionals to evaluate your plan's investment options and allocate your assets among themo Want to delegate the ongoing management of your workplace savings to Fidelity's investment professionalso Are willing to pay an annual advisory fee for ongoing professionalmanagement of your workplace savings accountWhat will the Service do for me and what will I be expected to do?Using your expected retirement age or your responses in your Personal Profile as aguide, the Service will take on many of the day-to-day investment decisions on yourbehalf. The Service will:o Determine the initial and ongoing investment mix in your plan account to align with a model portfolio that seeks to support growth and manage risko Change the investment mix of your future contributions to align with your model portfolio (note, however, that the Service does not control the amountof the contributions you choose to make to your retirement savings account)o Rebalance and reallocate your workplace savings plan account through the ups and downs of the marketo Provide confirmations showing transactions initiated by the Service on your behalfo Provide account statements detailing your holdings and investment mix, as well as any advisory fees paid from your account during the statement periodo Contact you annually to review your investment approach to make sure that it is still appropriate for your future goalsYou will continue to decide how much to contribute to your workplace savingsaccount, make loan or withdrawal decisions for your workplace savings account, andmake decisions about your company stock, if you have any. You may choose tocomplete your Personal Profile at any time, if you have not done so. We will use thisadditional information to provide a more comprehensive analysis of your workplacesavings account.Beyond this, you should update your information with us whenever a personal orfinancial change occurs.Can I contact someone about my managed account once I've signed up for this Service?Yes, you can speak to a dedicated Workplace Planning Consultant by calling 1-866-811-6041. These Consultants can provide you with the latest information on youraccount's investment holdings and offer general account assistance.How does a managed account work?It's easy to enroll. To get you started we assign a strategy based on the things weknow about you, either through your employer or your relationship with Fidelity: your age, your salary, and the value of your retirement accounts or pensions with Fidelity.We also look at your anticipated retirement age, and how you would like us toconsider your company stock, if you own any. If you prefer a more comprehensiveapproach, which is what we recommend, you can choose to answer a series ofquestions about your financial situation through your Personal Profile. We'll propose an investment approach based on your responses. Either way, we will use yourinformation to suggest an investment approach and a model portfolio designed for an investor like you, using the investments offered through your workplace savings plan.When you accept this investment approach, we will allocate your current plan account balance and invest your future contributions to align with the model portfolio. Theinvestments in your assigned model portfolio may be reallocated when market oreconomic conditions warrant, if there are changes to the characteristics of the model portfolio holdings, or if your plan makes changes to its investment options. If younotify us of changes in your situation, the Service may rebalance your account toreflect the change, if appropriate.Once a year, we ask you to review your responses in your Personal Profile to helpensure your current investment approach is still appropriate for your retirement goals.If you revise your responses, we'll review your investment approach and makechanges, if necessary.Currently, we take financial responsibility for certain charges incurred during theongoing management of your account, such as purchase fees and redemption fees.However, we reserve the right to pass on to your account these types of fees that are typically generated when exchanges into or out of an investment option in youraccount occur. You are responsible for any redemption fees, if applicable, that mayresult from the initial exchanges when you enroll in the Service or from any loans orwithdrawals you initiate. However, if the Service initiates a transaction in your planaccount that results in a redemption fee, this fee will be covered by the Service unless we have changed our practice of assuming financial responsibility for these costs. What are the benefits of a managed account?• A team consistently reviews your investment strategy, and proactively adjusts it through market ups and downs to stay aligned with your retirement goals.•Risk tolerance, how much you’ve saved, and how long your money will need to last are all taken into account.•Access to an exclusive dashboard to make any updates based on life events. track income estimates, view your progress toward retirement.Who will be managing my workplace savings account?Your Fidelity managed account will be managed by Strategic Advisers and FidelityPersonal and Workplace Advisors, both are registered investment advisers andFidelity Investments companies. For nearly 30 years, Strategic Advisers has provided professional investment management and financial planning and advice tools toFidelity's customers. Following asset allocation principles, along with deepquantitative and fundamental research, Strategic Advisers and Fidelity Personal and Workplace Advisors LLC's disciplined approach to investment management thatseeks risk-adjusted results over time—all designed to help give individuals andfamilies the reassurance they need.See the Terms and Conditions for the education and business backgrounds of theinvestment professionals who develop model portfolios.What is your Personal Profile?For a more comprehensive analysis of your situation, your Personal Profile guides you through a series of questions to determine your preferred investment style, howmuch you may need in retirement, your anticipated income sources, and yourretirement time frame. This information will help us determine an appropriateinvestment approach of stocks, bonds, and short-term investments for your assigned model portfolio.How is company stock considered?Investing a portion of your plan in company stock can play an important role inhelping you meet your long-term retirement goals. However, a large holding of asingle stock, including company stock, may create higher risk.Your company stock is not part of the assets that are managed by the Service. You can direct the Service to (1) attempt to offset the risks of your company stock in ourapproach, (2) ignore your company stock in our approach, or (3) sell all your company stock (unitized company stock fund only). You can also sell your company stockbefore enrolling or updating your plan. You may wish to consult a tax advisor before making a decision about selling your company stock.•Offset Company Stock: Offsetting the risk associated with company stock may result in a more conservative investment approach than if your stock holdingswere ignored. If you request that Fidelity attempt to "offset" the characteristics ofcompany stock in your account, they will assign you to a model portfolio thatstrives to offset the risks associated with owning company stock. If, as a result ofyour company stock position, your offset model portfolio consists exclusively ofbond funds and short-term investments, you should carefully consider whetherthis fee-based service is appropriate for you.•Ignore Company Stock: Ignoring the risk associated with company stock may result in a more aggressive investment approach than if your stock holdings wereoffset. If you request that Fidelity "ignore" the characteristics of company stock inyour account, they will assign you to a model portfolio that ignores the risksassociated with owning company stock.•Sell Company Stock: (Only available for company stock traded in a unitized stock fund). You can direct the Service to sell all of your company stock holdings in thisplan for you. You should carefully consider the tax and other implications ofselling company stock held within your account. You may wish to consult a taxadvisor before making a decision.For more information on how the Service treats company stock, please see the Terms and Conditions.Why am I unable to invest in BrokerageLink® when enrolled in Personalized Planning & Advice?If you are currently enrolled in Personalized Planning & Advice and your plan allowsall assets to be managed by this service, enrolling or depositing additional moniesinto BrokerageLink is not allowed. If you wish to enroll in BrokerageLink, or if yourplan allows, contribute more to BrokerageLink , you must first un-enroll fromPersonalized Planning & Advice.For more information on how to un-enroll before opening or contributing additional monies into your BrokerageLink account, please contact a service representative.How are BrokerageLink® assets considered by Personalized Planning & Advice?Personalized Planning & Advice does not manage the BrokerageLink® assets;however, the Service takes in to account the equity percentage of the BrokerageLink assets to determine an appropriate investment approach of stocks, bonds, and short-term investments for your assigned model portfolio.For more information on how the Service treats BrokerageLink assets, please contacta service representative.What does "Other" mean under "Not eligible for Personalized Planning & Advice" and how is it treated by the Personalized Planning & Advice?Your plan sponsor may elect to exclude certain investment options in your line-upfrom Personalized Planning & Advice. In these cases, the Service does notmanage those investment options; however, it does consider the equity percentage of these excluded investments to determine an appropriate investment approach of stocks, bonds, and short-term investments for your assigned model portfolio.These assets may also be restricted from exchange by plan rules. To exchange these assets, or determine which investment options have been excluded please contact a service representative.How can I access my account information?There are many ways to stay informed about your managed account.o You may view your account through NetBenefits®, virtually 24 hours a dayo You will automatically receive confirmations showing transactions initiated by the Service on your behalfo You will receive or may view online your account statement, detailing your holdings and investment mix, as well as any advisory fees paid from youraccount during the statement periodo You can speak with a Fidelity representative by calling 1-866-811-6041. These representatives can provide you with the latest information on your account'sinvestment holdings and offer general account assistance.What if my financial situation changes?Whenever there are changes to your personal or financial situation, you will need to update your information to keep your approach on the right track. Changes include:o Changing your anticipated retirement ageo Selling some of your assets outside of your managed accounto Receiving an initial or large contribution in company stocko Selling your company stock holdingso A significant change in your annual incomeYou can update your information online virtually 24 hours a day or by calling adedicated Fidelity representative at 1-866-811-6041.What if I need to take a loan or withdrawal from my plan account?You would follow the same procedures that your workplace savings plan currently has in place. If you take a partial withdrawal, your remaining assets will continue to bemanaged. If you take a full payout, you will no longer be enrolled in the Service. Inthis case, the Service will automatically deduct from your payout amount the prorated advisory fee based on the number of days you used the Service during the statement period, unless your plan sponsor is paying for the Service.What if I cancel the service?There is no cancellation fee should you decide to terminate the Service. You maycancel at any time by calling 1-866-811-6041. Unless you take action to change yourinvestment mix, your workplace savings account will remain invested in accordancewith the last investment mix determined by the Service and all additionalcontributions to your account will be invested in accordance with those allocations.You will receive a confirmation when you cancel the Service.We will automatically deduct a prorated advisory fee based on the number of daysyou used the Service during the statement period, unless your plan sponsor is paying for the Service. Sometimes a transaction that is in-process when you cancel results ina short-term trading fee after you cancel the Service--if that happens, you will beresponsible for paying this short-term trading fee.Can I make changes to my investments?Once you are enrolled in the Service, you cannot make changes to your investmentmix for the account(s) being managed. When you choose a managed account, youturn over the day-to-day management of your investments to a team of professionals.The Service determines the investment mix on your behalf and changes theinvestment mix for future contributions to align with your assigned model portfolio.What happens if I leave my current employer? Can I leave my money in the account?This depends on your plan rules. As long as your plan sponsor allows you to remain in the plan, you can continue to enjoy the benefits of Personalized Planning & Advice.How accurate do I need to be?While it's not necessary to be accurate down to the last penny, providing moredetailed, accurate information can help Fidelity develop a more comprehensiveproposal that more closely reflects your overall financial situation. For this reason, ithelps if you can be reasonably accurate.How is this different from a target date fund?While a target date fund is a good retirement savings solution for many workplaceinvestors, it may not be as comprehensive as Personalized Planning & Advice. With a target date fund, the investment mix is dictated primarily by the date associated with the fund, which is assumed to be at or near the investor's retirement date. So forinstance, a fund will have a certain prescribed mix of stocks, bonds, and moneymarket investments 20 years prior to the target date, a different prescribed mix 10years prior to the date, and so on, with the mix slightly changing until 15 years afterthe target date has passed. While these types of funds can help investors looking for an age-appropriate investment mix, they don't account for any data other than age.Personalized Planning & Advice is designed to be much more comprehensive,drawing on a range of personal and financial data, including (depending onhow many of the questions you answer), your age, your salary and plancontributions, the precise year you hope to retire, the number of years you'll needmoney, the amount you've saved in both retirement and non-retirement accounts,any income you may receive from things like Social Security or a pension, yourfeelings about investment risk, your investment knowledge and experience, and your overall financial picture.What if I expect this information to change in the future?In order to ensure that your investment mix is based on the most up-to-dateinformation, Fidelity recommends that you revisit this questionnaire annually, making adjustments to reflect your changing situation. You should also come back and revisit this tool if you've gone through a major life event, such as a marriage, or if theanswers to some of these questions have changed significantly. For instance, you may have increased the contributions you make to your workplace plan, or you may have changed the way you view investment risk. Keeping your information as current aspossible is the best way to ensure that you're maintaining an investment mix that'sappropriate for you.What if I have significant income not reflected here?If you have significant amounts of other income from a second job, a rental property you own, or a structured settlement, or if you think your financial situation is morecomplex than what's shown here, you can provide more detail in the questionnaire.It's been designed to go into greater depth and can help capture some of thesedetails.Do I have to answer every question?While it's not necessary to answer every question, it's important to remember thathaving more information about you can help Fidelity develop a more comprehensive proposal and an investment mix that more accurately reflects your unique financialsituation.Can I have my non-workplace accounts managed by this service?No. Personalized Planning & Advice is not available for personal accounts. Fidelitydoes however have other managed account options for personal accounts. For more information call 866-811-6041.Learn more about Personalized Planning & Advice at /PPA.Keep in mind that investing involves risk. The value of your investment will fluctuate over time and you may gain or lose money.Fidelity® Personalized Planning & Advice at Work is a service of Fidelity Personal and Workplace Advisors LLC and Strategic Advisers LLC. Both are registered investment advisers and Fidelity Investments companies and may be referred to as “Fidelity,” “we,” or “our” within. For more information, refer to the Terms and Conditions of the Program. When used herein, Fidelity Personalized Planning & Advice refers exclusively to Fidelity Personalized Planning & Advice at Work. This service provides advisory services for a fee.You may unenroll by calling a representative. There are no cancellation fees.Fidelity Brokerage Services LLC, Member NYSE, SIPC, 900 Salem Street, Smithfield, RI 02917© 2023 FMR LLC. All rights reserved.1083822.1.0。

SimpliFi HS Mix商品说明书

Website:/email:****************************PI -50546 V1Storage and stability:SimpliFi HS Mix is shipped on dry/blue ice. On arrival store at -20 °C for optimum stability. Repeated freeze/thaw cycles should be avoided. Thawing during transportation does not affect the product performance. Solutions should be mixed/equilibrated after each thawing to avoid phasing.Expiry:When stored under the recommended conditions and handled correctly, full activity of the kit is retained until the expiry date on the outer box label.Safety precautions:Read and understand the SDS (Safety Data Sheets) before handling the reagents. Hardcopies of the SDSs will be provided with the first shipment, thereafter they will be available upon request.Quality control:Meridian operates under ISO 13485 Management System. SimpliFi HS Mix and its components are extensively tested for activity, processivity, efficiency, heat activation, sensitivity, absence of nuclease contamination and absence of nucleic acid contamination.Notes:For research and further manufacturing use only.Store at –20 °CImportant considerations and PCR optimizationThe optimal conditions will vary from reaction to reaction and are dependent on the template/primers used.Mg 2+ concentration: The Mg 2+ concentration in the 2x mix is 4 mM (2 mM final concentration), this is the optimum concentration for SimpliFi HS Mix for most PCR reactions and should only be adjusted if necessary . Additional Mg (up to 4 mM in the final reaction) should be added in presence of more than 10% of whole blood.Primers: Forward and reverse primers are generally used at the final concentration of 0.2 - 0.6 μM each. As a starting point, we recommend using 0.4 μM final concentration (i.e. 20 pmol of eachprimer per 50 μL reaction volume). Too high a primer concentration can reduce the specificity of priming, resulting in non -specific products.When designing primers we recommend using primer -design soft-ware such as Primer3 (/primer3) or visual OMP TM (). Primers should have a melting temperature (Tm) of approximately 60 °C.Template: The amount of template in the reaction depends mainly on the type of DNA used. For templates with low structural complexity, such as plasmid DNA, we recommend using 50 pg - 10 ng DNA per 50 μL reaction volume. For eukaryotic genomic DNA, we recommend a starting amount of 200 ng DNA per 50 μL reaction, this can be varied between 5 ng - 500 ng. It is important to avoid using template re -suspended in EDTA -containing solutions (e.g. TE buffer) since EDTA chelates free Mg 2+.Multiplexing: For multiplex PCR we suggest using 55 °C as a starting annealing temperature. If further optimization is required we recommend using a temperature gradient to determine the optimal annealing temperature needed for the multiplex PCR. Since multiplex PCR generally requires a longer extension step, we suggest starting with a minimum of 90 s and increasing it if required.DescriptionSimpliFi HS Mix is a convenient ready -to -go 2x reaction mix combining the latest advances in buffer chemistry and PCR en-hancers and stabilizers, together with an aptamer -mediated hot -start polymerase, dNTPs and MgCl 2. It has been designed for highly reproducible, accurate assay results in the presence of inhibitors. The mix is optimized and ready -to -use, the user is simply required to add water, template and primers.The advanced buffer chemistry and enhancers in SimpliFi HS Mix have been developed for fast PCR and is designed for superior sensitivity and specificity. SimpliFi HS Mix has been developed to reduce GC bias, making it perfect for NGS library amplification. ComponentsFeatures• High fidelity coupled with high yield• Inhibitor -tolerant, amplifying of a broad range of targets • Low GC bias• Convenient pre -mixed, pre -optimized 2x solution • Reproducible resultsApplications• Ideal for crude samples such as blood • NGS library amplification • Multiplex PCR • Blunt -end cloningShipping:On Dry/Blue IceCatalog numbers: BIO -25060 : 100 x 50 μL reactions (2 x 1.25 mL) BIO -25061 : 500 x 50 μL reactions (10 x 1.25 mL) Batch No.: See vial Concentration:2xTroubleshooting GuideAssociated Products____________________________________________________________________________________________________________________________Bioline Reagents Ltd UNITED KINGDOM Tel: +44 (0)20 8830 5300 Fax: +44 (0)20 8452 2822Meridian Life Science Inc.USATel: +1 901 382 8716Fax: +1 901 382 0027Bioline GmbHGERMANYTel: +49 (0)337 168 1229Fax: +49 (0)3371 68 1244Bioline (Aust) Pty. LtdAUSTRALIATel: +61 (0)2 9209 4180Fax: +61 (0)2 9209 4763Technical SupportFor any technical enquiries, please contact our Technical Supportteam via email at: ********************************Website:/email:****************************。

Philips Fidelio B5 声音条说明书

Philips Fidelio SoundBar Soundbar speaker4.1 CH wireless surround sound Bluetooth ® aptX ,AAC and NFC 2 HDMI In & HDMI out ARC Spatial calibrationB5Stereo to surround sound in secondswith detachable speakersThe Philips Fidelio B5 soundbar creates surround sound using wireless detachable rear speakers and subwoofer. Audiophile drivers and spatial calibration ensures dynamic, balanced sound. Rears become independent speakers for music anywhere.Richer sound for watching TV and movies•Surround on Demand - detachable wireless surround speakers •Spatial calibration•Dolby Digital and Pro Logic II surround sound•Premium soft dome tweeters for Crystal Clear Sound •Near listening sound modeConnect and enjoy all your entertainment •Stand-alone portable Bluetooth speakers•Bluetooth ® (aptX ® and AAC) for wireless music streaming •Connect to HDMI x 2 for great picture and sound quality•One-Touch with NFC-enabled smartphones for Bluetooth pairing •HDMI ARC to access all the sound from your TV Designed to enhance your living space•Minimalistic chic design to suit a modern decor •Wireless subwoofer for clutter free placementSurround on DemandSubmerge your listening senses in superb surround sound whenever you want it. Simply detach the wireless surround speakers from the left and right main speaker units and place them at the rear of the room to getmultichannel surround sound experience with your games and movies. When the action is over, reattach the speakers for music and TV shows. Using proprietary wireless audio technology with surround speakers that operate on a unique frequency, there is no network interference. The result is acompletely wireless surround system with lossless audio quality for music and movies.Spatial calibrationAn advanced spatial calibration function developed by our unique innovation lab ensures the best possible balance of sound, regardless of where in a room rear speakers have been placed. When triggered, thisfunction uses a specially designed tonal signal to locate each of the rear speakers. It then uses this information to identify the best sound balance for this speaker placement and adjusts the sound of each speaker to match. This allows the listener to enjoy properly balancedsound even if speakers are placed outside of the ideal position.Premium soft dome tweetersWith Philips' Crystal Clear Sound, you will now be able to discern and appreciate every sound detail, the way it's meant to be heard and as the artist or director intended! Crystal Clear Sound reproduces the sound source - whether it's an action movie, a musical or a live performance - faithfully, precisely and without any distortion. Philips home theaters with Crystal Clear Sound keep audio processing to a minimum, preserving the purity of the original audio content. The result is greater sonic accuracy delivered to your ears.Bluetooth ® (aptX ®and AAC)Bluetooth ® (aptX ® and AAC) for wireless music steamingStand-alone portable speakersBoth of the Fidelio B5's detachable rear speakers feature an independent Bluetooth module, making them truly independent. This means they can be taken anywhere, in the home or out, for total freedom of musical enjoyment. Each portable speaker has a total power output of 8W, so they can deliverpowerful sound wherever they are. With two rear detachable speakers and a central sound bar unit, a single Fidelio B5 unit can bring music to three different places at the same time.NFC technologyPair Bluetooth devices easily with one-touch NFC (Near Field Communications) technology. Just tap the NFC enabledsmartphone or tablet on the NFC area of a speaker to turn the speaker on, startBluetooth pairing, and begin streaming music.HDMI x 2Enjoy larger-than-life 3D playback and crystal-clear 5.1 or 7.1 audio by simply plugging our player's audio HDMI x 2 output to the connection in your non-3D AV receiver.Dolby Digital and Pro Logic IIA built-in Dolby Digital decoder eliminates the need for an external decoder by processing all six channels of audio information to provide a surround sound experience and anastoundingly natural sense of ambience and dynamic realism. Dolby Pro Logic II provides five channels of surround processing from anystereo source.Issue date 2022-04-24 Version: 4.5.412 NC: 8670 001 20329 EAN: 04 89518 56041 52© 2022 Koninklijke Philips N.V.All Rights reserved.Specifications are subject to change without notice. Trademarks are the property of Koninklijke Philips N.V. or their respective owners.Product dimensions•Product dimensions (W x H x D):103.4 x 7.17 x 15.55 cm•Weight: 4.49 kgPackaging dimensions•Packaging dimensions (W x H x D):75.7 x 35.7 x 25.2 cm•Nett weight: 10.2 kg•Gross weight: 12.58 kg•Tare weight: 2.38 kg•EAN: 48 95185 60415 2•Number of products included: 1•Packaging type: Carton•Type of shelf placement: LayingOuter Carton•Outer carton (L x W x H): 76.5 x 26 x 37.1 cm •Nett weight: 10.2 kg•Gross weight: 13.28 kg•Tare weight: 3.08 kg•EAN: 48 95185 60415 2•Number of consumer packagings: 1Sound•Sound Enhancement: Surround on demand, Night Mode, Treble and Bass Control, Audio sync, Auto Volume Leveler, Movie, Music, Near Listening, Voice•Sound System: Dolby Digital 5.1, DTS Digital Surround•Subwoofer output power: 90•Total Power RMS @ 30% THD: 210 W •Speaker output power: 120Loudspeakers•Drivers per Surround speaker: 1 X 3" full range speaker•Subwoofer driver: 1 x 6.5" woofer •Subwoofer freq range: 20-150 (-3dB) Hz •Subwoofer impedance: 4 ohm •Subwoofer type: Wireless subwoofer, Bass reflex system•System: 2.1/4.1 system•Drivers per Soundbar: 2 X 3", 2X 1" soft dome tweetersConnectivity•Rear Connections: Digital coaxial in, Digital optical in, HDMI 1.4 output (ARC), HDMI IN 1, HDMI IN 2, Audio in 3.5 mm jack•Wireless connections: Bluetooth APT-X and AAC Convenience•EasyLink (HDMI-CEC): Audio Return Channel, Automatic audio input mapping, One touch standby, Remote Control-Passthrough•HDMI Features: Audio Return Channel (ARC), Content TypePower•Power supply: 110-240V, 50Hz•Standby power consumption: <0.5 W Sustainability•Packaging: 80% recycled corrugated board, Soya based inkDimensions•Main Unit (W x H x D): 1035 x 70 x 156 mm •Main Unit Weight: 4.18 kg•Subwoofer (W x H x D): 200 x 510 X 200 mm •Subwoofer Weight: 5 kg•Weight incl. Packaging: 16 kg•Packaging (W x H x D): 765 x 260 x 371 mm Accessories•Included accessories: 2 x AAA Batteries, Power cord, Remote Control, Safety & Legal Leaflet, Trademarks Sheet, User Manual, World Wide Warranty leaflet, Wall mount bracket。

AgeI限制性内切酶使用说明书

H om e ›P roducts ›AgeIAgeIProduct Information FAQs Protocols Other Tools & Resources Related ProductsDescriptionProperties and UsageQuality Control This enzyme has transitioned to an improved new buffer system. Visit for further details.The new and current Double Digest Finder and current Activity/Performance Chart for the CutSmart buffer system are available. The previous version of the Double Digest Finder , as well as the previous Version of Activity/Performance Chart that use the former buffer system, are still available for your convenience.AgeI has High Fidelity (HF) and RE-Mix Master Mix versions available.IsoschizomersCatalog #Size Concentration Price Qty R0552S300 units 5,000 units/ml $67.001R0552L 1,500 units 5,000 units/ml $268.001Categories: Restriction Endonucleases: AApplications: Restriction Enzyme DigestionDescriptionReaction Conditions1X NEBuffer 1.1Incubate at 37°C1X NEBuffer 1.1:10 mM Bis-Tris-Propane-HCl10 mM MgCl2100 μg/ml BSApH 7 @ 25°CActivity in NEBuffersNEBuffer 1.1: 100%NEBuffer 2.1: 75%NEBuffer 3.1: 25%CutSmart™ Buffer: 75%Diluent CompatibilityDiluent CStorage Temperature-20°CStorage Conditions10 mM Tris-HCl250 mM NaCl1 mM DTT0.1 mM EDTA200 μg/ml BSA50% Glycerol0.15% Triton® X-100pH 7.4 @ 25°CHeat Inactivation65°C for 20 minMethylation Sensitivitydam methylation: Not Sensitiv edcm methylation: Not Sensitiv eCpG Methylation: BlockedActivity at Temperature@37°C: 100%Quality ControlQuality Control AssaysThe following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Indiv idual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page. Blue-White Screening (Terminal Integrity):A sample of DNA v ector linearized with a 10-fold excess of a restriction endonuclease, religated and transformed into an E. coli strain expressing the LacZ beta fragment gene results in less than 1% white colonies.Exonuclease Activity (Radioactivity Release):The product is tested in a reaction containing a radiolabeled mixture of single and double-stranded DNA. After incubation for 4 hours the exonuclease activ ity is determined by the % release of radioactiv e nucleotides.Ligation and Recutting (Terminal Integrity):After an ov er-digestion of DNA with a restriction endonuclease the percentage of the DNA fragments ligated with T4 DNA ligase andthe percentage that can be recut are determined by agarose gel electrophoresis.Non-Specific DNase Activity (16 hour):The product is tested for non-specific nuclease degradation in a reaction containing a DNA substrate. After incubation for 16 hours there is no detectable degradation of the DNA substrate as determined by agarose gel electrophoresis.Notes1. Star activ ity may result from extended digestion.Legal and DisclaimersPatentsNew England Biolabs, Inc.U.S. Patent No. 7,517,671Supporting DocumentsCertificate of AnalysisThe Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality control's for an indiv idual lot. The following file naming structure is used to name these document files: [Product Name]COA.R0552SLCoA-Lot0431302Material Safety DatasheetsThe following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name]MSDS. For international v ersions please contact us at info@ .AgeI MSDSNEBuffer 1.1 MSDSDatacardsThe Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@ or fill out the Technical Support Form for appropriate document.R0552Datasheet-Lot0431205R0552Datasheet-Lot04312091. What factors inhibit AgeI?2. Are extended digests of AgeI recommended?3. What is the reaction temperature of AgeI?4. My enzyme is no longer Time-Sav er™ qualified. What happened?5. What effect does BSA hav e on the performance of NEB’s restriction enzymes when included in the new buffers?6. How can I access the old NEBuffer Activ ity Chart?7. How can I access the old Double Digest Finder?1. Optimizing Restriction Endonuclease ReactionsSelection Tools Usage Guidelines & TipsTroubleshooting Guides Interactiv e ToolsSelection ToolsAlphabetized List of Recognition SpecificitiesCompatible Cohesiv e EndsCross Index of Recognition SequencesDam-Dcm and CpG MethylationFrequencies of Restriction SitesIsoschizomersRecleav able Filled-in 5' Ov erhangsTime-Sav er™ Qualified EnzymesWhy Choose Recombinant Enzymes?Usage Guidelines & TipsActiv ity at 37°C for Restriction Enzymes with Alternate Incubation TemperaturesActiv ity of Restriction Enzymes in a Taq or Phusion® PCR MixActiv ity of Restriction Enzymes in a Vent R® PCR MixCleav age Close to the End of DNA FragmentsCleav age of Supercoiled DNADigestion of Agarose-Embedded DNA: Info for Specific EnzymesDouble DigestsEffects of CpG Methylation on Restriction Enzyme Cleav ageHeat Inactiv ationMegabase MappingNEBuffer Activ ity/Performance Chart with Restriction Enzymes Optimizing Restriction Endonuclease Reactions Restriction Endonucleases - Surv iv al in a Reaction Restriction Enzyme Diluent Buffer CompatibilitySingle Letter CodesStar Activ ityTraditional Cloning Quick GuideTroubleshooting GuidesRestriction Enzyme Troubleshooting GuideInteractive ToolsDNA Sequences and Maps ToolDouble Digest FinderEnzyme FinderCompanion ProductsAgeI-HF™AgeI-HF® RE-Mix®Materials Sold SeparatelyNEBuffer 1.1。

常用载体构建说明书

常用载体构建说明书步骤：一、基本耗材准备（抗生素、LB液体培养基、LB固体培养基、离心管、枪头、三角瓶）二、制备感受态大肠杆菌三、设计引物四、Pcr扩增五、Pcr产物检测六、Pcr产物回收七、双酶切pcr产物和质粒八、酶切产物回收九、目的基因与载体连接十、转化感受态大肠杆菌十一、单克隆检测十二、测序比对十三、提质粒十四、酶切验证十五、转化感受态农杆菌十六、单克隆检测十七、侵染液配制一、基本耗材准备1、抗生素的制备（抗生素为索来宝公司）常用抗生素Kan（卡那）Amp（氨苄）Rif（利福平）母液浓度：Kan 50mg/ml Amp 100 mg/ml Rif 100 mg/ml工作浓度：Kan 50ng/ul Amp 100 ng/ul Rif 100 ng/ul举例：称取kan固体1g 于注射器中，加入20ml ddH2O，溶解后用过滤器注入灭过菌的离心管中，-20度保存，使用比例1:1000注意：Rif溶解时加入DMSO2、培养基的配制LB液体培养基1000ml 200ml牛肉膏5g 1g蛋白胨10g 2g氯化钠10g 2gLB固体培养基1000ml 200ml牛肉膏蛋白胨氯化钠琼脂粉5g10g10g15g1g2g2g3g120℃高温高压灭菌15-20min，固体培养基冷却后加入相应抗性，加入比例1:1000，然后把培养基倒90cc的培养皿中，一般情况下一个培养皿可倒20ml培养基。

3、离心管、枪头、三角瓶0.2ml、1.5ml、2.0ml离心管各200-500个，50ml离心管2个10ul、200ul、1000ul 枪头各2盒50ml、100ml、250ml 、500ml三角瓶各2个去离子水或双蒸水500ml二、制备感受态大肠杆菌（所用超级感受态细胞制备试剂盒购于上海生工）BT Media 培养基制备：1支BT Media加50 ml蒸馏水配制，放入250ml三角瓶，高压灭菌即可准备工作：将BT Buffer A和BT Buffer B 放冰上遇冷，遇冷低温离心机至4℃1、用超低温冰箱中保存的菌种在LB平板上进行划线，置37℃培养箱中静置培养12-16h待菌落生长到1-2mm大小。

Uniden PRO 520XL 说明书

PRO 520XLProfessionalMobile CB RadioPrinted in China.UTZZ01363EZ(0)Welcome to the world of Citizens Band radio communications. Your Uniden radio is an advanced mobile radio designed for use in the Citizens Band (CB) Radio Service. It will operate on any of the 40 AM frequencies authorized by the Federal Communications Commission (FCC). Your Radio features a superheterodyne circuit with PHASE LOCKED LOOP techniques to assure precise frequency control. WARNINGThis radio has been type accepted and certified by the FCC for CB Radio Service operation. Any adjustments or alterations which might alter the performance of the transceiver or the method of determining frequency are strictly prohibited.If you replace or substitute any unique parts (including crystals, transistors, IC’s, regulator diodes, etc.) with parts other than those recommended by Uniden, you may be in violation of FCC technical regulations (in Part 95) or type acceptance requirements (in Part 2) of the FCC rules.CB license requirementU.S. users no longer need to apply for a license to use a CB radio. However, CB radio is still considered a “license by rule” service. This means that, while the FCC permits CB station operation without station identification, station operators are still required to comply with the Communications Act and with the rules of CB Radio Service. (For more information on U.S. radio rules, visit the FCC website at .) Uniden contact informationYou can get answers 24/7 at our website: .If You Contact Uniden’s Phonehave a question or problem Customer Care Line*800-297-1023 need a part or accessory Parts Department*800-554-3988need special assistancedue to a disability Accessibility Help Line 800-874-9314 (voice or TTY)*During regular business hours, Central Standard Time. Visit our website for detailed business hours.If any of these items are missing or damaged, immediately contact your place of purchase or Uniden’s Parts Department.PRO520 XL Mobile radioMicrophone and hookMounting bracket, knobs, and mounting hardware Other printed materialsNOTE: You must use a CB antenna (sold separately) with this radio.Front1. PA SwitchActivatesthe PublicAddressConnectingexternalspeakers).2. ANLSwitchActivatesthe Automatic Noise Limiter feature, which reduces backgroundnoise from the car engine and other sources. Move this switch up to turn on the ANL feature.3. CH9 SwitchSwitches the radio over to channel 9.►INCLUDED IN YOUR PACKAGEPARTS OF THE RADIO4. TX LED Indicates when the radio is transmitting.5. S/RF (Signal/RF) Meter Shows the strength of the received signal or the RF output.6. Channel Indicator Displays the channel currently in use.7. Microphone Jack Connects to the included detachable electret microphone.8. Volume Control Powers on the radio and adjusts the volume.9 Squelch knob Adjusts the level at which the radio squelches or suppresses weak radio signals.10. RF Gain Knob Adjusts the level of the signal reception.11. Channel Selector Selects which channel the radio will transmit or receive on. Back12. Model labelnumber.13.Antenna Connector14. See 15. External Speaker Jack See Connecting external speakers , below.16. 12 Volt DC Power Cord17. Fuse casing Microphone18 Push-to-talk (PTT) buttonNOTE: Make sure you have read and understood part 95 of the FCC rules and regulations before using the transmitter.Connect the microphoneAlign the microphone connector with the jack on the front of the radio. Push the connector in firmly and secure it with the locking screw.Connect the powerYou can connect the radio to any standard 12 volt DC power source, with either a positive or negative ground. If you don’t know whether your power supply uses a positive or negative ground, consult the manual for your power supply or contact the manufacturer.WARNING! DO NOT connect this equipment to a power supply if you are not absolutely certain of the grounding type!Make sure your power supply is turned off.For power supplies with a NEGATIVE ground, connect the RED lead on the radio to the power supply’s POSITIVE (+) pole, and connect the BLACK lead to the power supply’s NEGATIVE (–) pole or to a neutral ground such as the chassis. OR For power supplies with a POSITIVE ground, connect the BLACK lead on the radio to the power supply’s NEGATIVE (–) pole, and connect the RED lead to a neutral ground such as the chassis.Turn on your power supply.Turn the radio’s Volume knob clockwise to power on the radio.Installing the mounting bracketWhen choosing the location for the radio’s mounting bracket, keep the following things in mind:Pick a location that does not block your view, interfere with your vehicle’s controls, or hinder your driving.Make sure the radio and microphone are not in front of an airbag.Pick a solid surface that can support the weight of the bracket and the radio.Make sure there’s enough room. (You may want to put the radio in the bracket when you’re choosing where to install the bracket.) Once you choose the location, use the included, self-tapping screws to attach the mounting bracket and the microphone bracket to your vehicle (you don’t have to drill holes). Slide the radio into the bracket and use the included knobs to hold it at the preferred angle.1)2)3)4)►►►►Connecting an external antennaWARNING! The antenna used for this radio must be installed at least 8 inches (20 cm) away from all persons. The antenna must not be collocated or used with any other antenna or transmitter.CAUTION: Never operate your radio with no antenna or with a damaged antenna cable. This can damage the radio.You will need to purchase an antenna to operate the radio. There are two basic types of mobile CB antennas--full-length whips and loaded whips--with a wide variety of mounts to suit different vehicle locations.Choose an antenna that matches the specifications of this radio.Follow the manufacturer’s installation instructions carefully.Tune your antenna using a Standing-Wave Ratio (SWR) meter: set the radio to channel 20, and adjust the antenna until the SWR is as close as 1:1 as possible.CAUTION: Make sure the SWR is less than 2:1 before using the radio. An SWR higher than 2:1 can damage the transmitter.Your Uniden dealer can help you select the antenna that is best for your needs. Consult the specifications in the back of this manual for detailed transmitter and antenna information.Connecting external speakersYour radio supports two external speakers for remote monitoring and public address features. External speakers are sold separately.To prevent feedback, direct all speakers away from the microphone.External monitor PA speaker Function Remote receiver monitoring orsubstitution for internal speaker Public address broadcasts Impedance8 Ohms8 OhmsRating7 Watts7 WattsConnect to EXT.SP. jack (⅛-in/3.5 mm)PA.SP. jack (⅛-in/3.5 mm)Notes The internal speaker is disabledwhen an external speaker isconnected.You must have a PA speaker connected to use the PAfeature.►►►NOTE: FCC rules reserve Channel 9 for motorist assistance and other emergency communications. Channel 9 should only be used in situations where there is immediate danger to the safety of individuals or the protection of property.Make sure you read and understood part 95 of the FCC rules and regulations before using the transmitter.Turning theradio on Turn the Volume knob clockwise until the display backlight comes on.Turning the radio off Turn the Volume knob counter-clockwise until it clicks and the display backlight turns off.Selecting a channel Turn the Channel knob clockwise to move up the channel list. Turn it counter-clockwise to move down the channel list.Changing the volume Turn the Volume knob clockwise to increase the volume; turn it counter-clockwise to decrease the volume.TransmittingTune the radio to the channel you want to transmit on, and listen to make sure the channel is clear.Press and hold the PTT button.Hold the microphone about 2 inches away from your mouth and speak in a normal voice.Release the button to listen for a response.––––Using the PA featureMove the PA switch up; this turns off the CB radio and activates the PA speaker.On the microphone, press and hold the PTT button.Hold the microphone about 2 inches away from your mouth and speak in a normal voice. Use the Volume knob to adjust the volume of the PA speaker.When you’re finished broadcasting, release the button and move the PA switch back down.––––Adjusting the squelch levelTo filter out weaker signals and background noise, turn the knob clockwise to increase the squelch level.To decrease the squelch level so you can hear weaker radio signals, turn the knob counter-clockwise.––Adjusting the RF gainTurn the knob clockwise to boost the reception of weak signals, or counter-clockwise to reduce the reception of strong signals.In areas where strong signals cause noise and distortion, reduce the RF gain (turn the knob counter-clockwise).In areas where weaker signals are difficult to hear, increase the RF gain (turn the knob clockwise).–––Using the Instant Channel 9 featureMove the CH9 switch up; the radio automatically tunes to channel 9 and disables the channel selector.Move the switch back down to return to the previous channel and enable the channel selector.––Maintenance Every six to twelve months, check to make sure that...The Voltage Standing Wave Ratio (VSWR) is less than 2:1.All electrical connections are secure and free of corrosion.The antenna cable shows no wear or damage.All mounting screws are securely fastened.4. Inspect all screws and other mounting hardware.If your radio is not performing to your expectations, please try these simple steps. If these steps don’t solve your problem, call the Uniden Customer Service Center.Problem:Things to try:Radio won’t turn on (no power) 1. Check the radio’s power cord and all connections.2. Check the fuse in the radio’s power cord.3. Check your vehicle’s electrical system.Poor reception 1. Adjust the squelch level.2. Check the antenna, cable and connectors.3. Check operation mode of the radio.Background noise 1. Turn on the automatic noise limiter.2. Adjust the squelch level.Weak transmission 1. Check the antenna, cable and connectors.2. Check the antenna grounding.3. Check for corrosion on the connectors.Service and repair informationTechnical information, diagrams and charts will be provided upon request.Service, repair, or alignment should only be attempted by a qualified and/or licensed radio technician.When ordering parts, it is important to specify the correct model number of this radio.It is the user’s responsibility to make sure the radio is operating in accordance with the FCC Citizens Radio Service regulations at all times.►►►►►►►►G eneralChannels40 AMFrequency Range26.965 to 27.405 MHzFrequency Control Phase Locked Loop (PLL) synthesizerFrequency Tolerance±0.005%Operating Temperature-30ºC to +50ºCMicrophone Plug in type, dynamicInput Voltage13.8 V DC nom. (+ or - ground)Current Drain TX full mod., 1.7ARX with max. audio output, 1.7ASize4-1/2” W x 6-3/4” D x 1-3/8” HWeight 1 lb. 10 oz.Antenna Connector UHF, SO-239LED Meter Indicates relative RF output and received signal strength T ransmiTTerPower Output 4 WattsModulation Class B amplitude modulationFreq. Response300-2500 HzOutput Impedance50 ohms, unbalancedr eceiverSensitivity0.5μV for 10 dB; (S + N) /N typical (limit 1.0μV) Selectivity 6 dB @ 7kHz, 70 dB @ 10kHz typicalImage Rejection80 dB typicalI.F. Frequency Double Conversion Superheterodyne1st 10.692 MHz2nd 450 kHzRF Gain Control Adjustable for optimum receptionAutomatic Gain Control (AGC)less than 10dB change in audio output for inputs from 10 to 50,000 microvoltsSquelch Adjustable; threshold less than1μVAudio Output Power7 watts max. into 8 ohmsFreq. Response300 to 2000 HzDistortion less than 10% at 4 watts, 1000HzInternal Speaker16 ohms, 3 watts roundSpecifications and features are subject to change without notice.WARRANTOR: UNIDEN AMERICA CORPORATION (“Uniden”)ELEMENTS OF WARRANTY: Uniden warrants, for two years, to the original retail owner, this Uniden Product to be free from defects in materials and craftsmanship with only the limitations or exclusions set out below.WARRANTY DURATION: This warranty to the original user shall terminate and beof no further effect 24 months after the date of original retail sale. The warranty is invalid if the Product is (A) damaged or not maintained as reasonable or necessary, (B) modified, altered, or used as part of any conversion kits, subassemblies, or any configurations not sold by Uniden, (C) improperly installed, (D) serviced or repaired by someone other than an authorized Uniden service center for a defect or malfunction covered by this warranty, (E) used in any conjunction with equipment or parts or as part of any system not manufactured by Uniden, or (F) installed or programmed by anyone other than as detailed by the owner’s manual for this product.STATEMENT OF REMEDY: In the event that the product does not conform to this warranty at any time while this warranty is in effect, warrantor will either, at its option, repair or replace the defective unit and return it to you without charge for parts, service, or any other cost (except shipping and handling) incurred by warrantor orits representatives in connection with the performance of this warranty. Warrantor,at its option, may replace the unit with a new or refurbished unit. THE LIMITED WARRANTY SET FORTH ABOVE IS THE SOLE AND ENTIRE WARRANTY PERTAINING TO THE PRODUCT AND IS IN LIEU OF AND EXCLUDES ALL OTHER WARRANTIES OF ANY NATURE WHATSOEVER, WHETHER EXPRESS, IMPLIED OR ARISING BY OPERATION OF LAW, INCLUDING, BUT NOT LIMITED TO ANY IMPLIED WARRANTIES OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE. THIS WARRANTY DOES NOT COVER OR PROVIDE FOR THE REIMBURSEMENT OR PAYMENT OF INCIDENTAL OR CONSEQUENTIAL DAMAGES. Some states do not allow this exclusion or limitation of incidental or consequential damages so the above limitation or exclusion may not apply to you. LEGAL REMEDIES: This warranty gives you specific legal rights, and you may also have other rights which vary from state to state. This warranty is void outside the United States of America.PROCEDURE FOR OBTAINING PERFORMANCE OF WARRANTY: If, after following the instructions in the owner’s manual you are certain that the Product is defective, pack the Product carefully (preferably in its original packaging). The Product should include all parts and accessories originally packaged with the Product. Include evidence of original purchase and a note describing the defect that has caused youto return it. The product should be shipped freight prepaid, by traceable means, to warrantor at:Uniden America CorporationParts and Service Division4700 Amon Carter BlvdFort Worth, TX 76155。

Soundstream DX5 五通道2路电子分频器说明书

Date of Install:
DESIGN FEATURES
This unit has been equipped with separate front and rear inputs to
preserve use of the head unit fading control. The subwoofer outputs’
unique tracking control allows the user to select constant bass or fadable
bass depending on the specific installation.
The low frequency section of the DX5 is controlled by a precision filter. Thistechnologyallowsveryaccuratetuningwitha minimumofelectronic parts in the chain. This filter affords a steep 24dB per octave slope which tightly limits the subwoofer to its intended bandwidth. The high pass, staggerable front and rear outputs employ a totally transparent 6dB per octave slope.
Location crnd Mounting
The DX5 is compact and generates virtually no heat. It can be located almost anywhere within the passenger compartment or trunk. Do not to attempt to install the DXห้องสมุดไป่ตู้ in th e engine compartment or in any outside

利安森Audio神经网络用户指南说明书

E R G U I| iiiREAD THIS FIRST DO NOT INCORRECTLY CONNECT MIC PREAMPS TO THE LIAISON!To avoid damaging your hard won gear and voiding your warranty, follow the simple steps below.•Do not connect gear equipped with phantom power- such as mic preamps and channel strips - to the insert loops in your Liaison. Activating the phantom power will damage the electronics in the monitor and parallel processing paths and void the warranty instantly!It may also damage other outboard gear that is parked down-stream of the offending piece by applying the phantom power to the inputs.A mic pre output as the source connected to either Buss A or BussB Input is perfectly OK -phantom power is applied to the INPUT pins of a mic pre.Correct Path: Mic > Mic Pre Input > Mic Pre Line Out > Liaison Buss Input•If you cannot follow the first rule and choose to connect a mic preamp to the Liaison inserts, DO NOT engage the phantom power.The Dangerous Music Teamiii | DANGEROUS MUSICTABLE OF CONTENTSOverview 1Liaison Rear Panel Features & Benefits 1Liaison Front Panel Features, Benefits & Connection 2Liaison Application Notes 10-16Specifications 17Warranty 18| iVTHANK YOUThank you for choosing products from the exciting and innovative line of Dangerous Music recording equipment. Many years of reliable service can be expected from our gear. This is made possible through careful design, construction, and component choices by recording industry veterans.The Dangerous Music Liaison is designed to increase productivity and improve the creative process by unleashing the potential of your outboard gear arsenal. There are comprehensive tips and application notes in this user’s guide, so don’t skip the read.If you have any suggestions for future products, feel free to forward them!ABOUT DANGEROUS MUSIC“Audio Integrity: non-negotiable.” This is the credo upon which Dangerous Music is founded. Conceived and designed by end users—not by engineers in lab coats- the results are products that resurrect dynamic range, punch, intelligibility and emotion. These high-fidelity, uncompromising signal paths are achieved by harnessing over 20 years of Chris Muth’s design wizardry; creator of infamous custom black boxes for world class facilities like Hit Factory, Masterdisk, Absolute Audio and Sterling Sound Mastering. Then the critical listening skills of golden-eared luminaries are applied, with the result being musical tools that fulfill the actual needs of today’s flexible computer based studio.LIAISON | 1 OVERVIEWTo survive in this modern day audio jungle, the contemporary studio must be flexible and agile; a facility that can adapt to any workflow, effortlessly. Tracking, mixing and mastering services oftentimes must be performed under one roof, utilizing every piece of outboard gear available.The Dangerous Liaison realizes this dream by providing unprecedented access to your outboard equipment. Connect up to six of your favorite units to the Liaison (more if you daisy chain, leverage a patchbay or add a Dangerous Master). Instantly audition any device or combination, change the order, dial in some parallel processing and then store these customized signal paths as presets to use again, on demand, anytime. True hard-wire bypass relays remove the gear entirely from the signal path; this includes outboard that lacks an integrated bypass or true hard-wire bypass. Mastering grade components switch instantly and silently, providing true A/B comparisons without coloration or latency. Pre and post-processing monitor outputs make it simple to audition between the original source and the treated product.Unpack this audio traffic tower and celebrate! Inside the box you’ll find the Liaison and a standard 3 Pin IEC power cable and the manual you’re reading now.Note: We recommend reading the entire manual. This box is deep and you just spent a pile of cash, so why just wade in the shallow end?Liaison: Rear Panel Connections[rear panel inputs and outputs pictured above]2 | DANGEROUS MUSIC1. INPUTS & OUTPUTSFeature: Premium grade XLR Connectors.Benefit: More surface area for happy electrons to migrate through and no off shore, cheap look alike connectors that “almost” fit.Note: All connections wired pin 2 hot as per AES standards.Feature: DB25 Connector.Benefit: Screws in and locks, plus we ran out of room on the rear of the unit for 4 more stereo outputs!Tip: ,Cables DO make a difference. Ask for Mogami 2534 interconnects.2. A/C CABLE & SWITCHING[rear panel power section pictured above]Feature: Three pin IEC cable.Benefit: Simple to replace if lost.Feature: Selectable voltage.Benefit: Travel with the Liaison everywhere in the world, connecting the ultimate collection of gear and making friends.Operation: Please review the mains voltage in the red window before plugging in the unit or electrifying events may transpire. To change it, use a screwdriver to gently pry the mains inlet module’s cover open and flip the fuse block around so the correctvoltage appears in the window.Tip: ,The power switch is on the inlet module. Seat the AC cable completely and use properly grounded power mains for safe and quiet operation.Tip: ,If the Liaison will not power on and the power is known to be good, check the fuses under the inlet module’s cover. Use 500 milliamp fast blow for 120V (America) and 250 milliamp fast blow for 240V (Europe).LIAISON | 33. INSERTS: INPUTS & OUTPUTS[rear panel insert section pictured above]Feature: Connect outboard equipment here (i.e. compressor, limiter, eq, reverb...)Benefit: Individually selectable via the front panel of the unit (more to follow on this).Feature: Separate send and return cables, each on XLR.Benefit: Standard issue, readily available cables.Tip: ,The ability to loop multiple pieces of gear on a single insert if desired (i.e. comp and limiter back to back).4. BUSS A & B: INPUTS & OUTPUTS[rear panel buss section pictured above]Feature: Individual access in and out to both buss a & buss bBenefit: Leverage all your outboard gear twice, on two discrete buss paths. For example, take the output of your favorite mic preamp and send it to buss a. Then send the signal into your session. Meanwhile, connect the output of your mix tobuss b. Now all of your gear is accessible all the time for tracking, mixing and mastering.Tip: ,: Take the output of of buss a and connect it to the input of buss b to make one contiguous chain. This will allow outboard that is connected later, to appear earlier. For example... the BAX EQ is in insert1 and the Variable Mu™4 | DANGEROUS MUSICLimiter is in insert 5. Selecting buss a’s insert5 and buss b ’s insert 1, the Variable Mu™ will now receive audio before the BAX EQ.Operation: Press insert button 5 on buss a and insert button 1 on buss b .Tip: , There is an additional insert, insert 6, located on the front panel.5. MONITOR OUTPUTS[rear panel monitor outputsectionpictured above ]Feature: 4 stereo outputs that provide input and output monitoring of each Buss.Benefit: Listen before the outboard gear of buss a , or after, or before the outboard gear of buss b , or after to determinewhether the fruits of you labor are helping or hindering your efforts toward audio nirvana.Tip: , When used in stand alone mode, you may monitor buss a input (preprocessing) and buss b output (post processing)Tip: , monitor outs are tapped off the busses (pre buss a , post buss a , pre buss b , post buss b) and do not affect the main bus inputs or outputs. (See diagram below and page 10 for pinout)MONITOR OUT 1LIAISON | 5 Liaison: Front Panel Features & Benefits1. ADDITIONAL INSERT 6[additional insert six pictured above]Feature: Connect additional outboard equipment here (i.e. client’s favorite, rented, begged, borrowed or...)Benefit: Individually selectable via the front panel of the unit (more to follow on this).Feature: Separate send and return cables, each on XLR.Benefit: Standard issue, readily available cables.Tip: ,The ability to loop multiple pieces of gear on a single insert if desired (i.e. comp and limiter back to back).2. DEVICE INSERTS TO BUSS A/B[device inserts and busses pictured above]6 | DANGEROUS MUSICFeature: Instantly and silently switch between outboard processors.Benefit: The human ear has a remarkably fleeting memory. Echoic memory studies have resulted in up to 4 seconds of retention. Unfortunately these are based on vocal cues (random word sequences) vs. audio auditions. The latter haveresulted in only up to 1000 milliseconds. That means no time for a patchbay or clumsy manual jack swaps. The Liaisonprovides true, instantaneous comparisons with no disconcerting pops and thuds.Feature: True Hard-Wire Bypass Relays.Benefit: When an insert is in bypass, the outboard unit is completely out of the circuit. This is the same as strapping an incredibly short cable between the insert points- all circuitry is avoided.Tip: ,Even if the equipment itself does not have a True Hard-Wire Bypass, the Liaison provides it.Feature: Color coded: Green = Engaged; White = BypassedBenefit: Visual confirmation of equipment activated.3. BUSS A & BFeature: Apply your outboard gear on two completely discrete stereo busses.Benefit: Leverage all your outboard gear twice, on two discrete buss paths. For example, take the output of your favorite mic preamp and send it to buss a. Then send the signal into your session. Meanwhile, connect the output of your mix tobuss b. Now all of your gear is accessible all the time for tracking, mixing and mastering.Operation: Both Busses can be provided with input individually. Their respective outputs must be wired to the destination.Tip: ,: Take the output of of buss a and connect it to the input of buss b to make one contiguous chain. This will allow outboard that is connected later, to appear earlier. For example... the BAX EQ is in insert1 and the Variable Mu™Limiter is in insert5. Selecting buss a’s insert5 and buss b’s insert1, the Variable Mu™ will now receive audio beforethe BAX EQ.4. THE FLIP BUTTONS[flip buttons pictured above]LIAISON | 7Feature: For both buss a and buss b, the flip buttons reverse the position of the inserts in pairs.Benefit: For example, to answer the age old mastering question of “what should come first, my equalizer or limiter?” let’s say the Variable Mu™ limiter is in insert1 and the BAX EQ is in insert2. To audition them in reverse order, simply hit theflip1-2 switch. Now you are monitoring the BAX EQ into the Variable Mu™ limiter.Tip: ,: Want to take advantage of parallel processing for the Variable Mu™ limiter in the above example? Since the parallel processing loop affects whatever processor is positioned in the even numbered inserts (i.e. 2, 4 and 6), and the VariableMu™ limiter is connected to insert1, simply reverse the order of the inserts with the flip1-2 switch and take advantageof actions like the blending of extreme limiting or compression. Now the insert1 button will blink slowly, indicating it’sactivation in the parallel processing loop.Tip: ,: The flip buttons affect both buss a and buss b simultaneously.5. THE PARALLEL PROCESSING LOOP BUTTONS[parallel processing loop buttons pictured above]Feature: parallel processing loop for the outboard equipment positioned in the even numbered inserts (i.e. 2, 4 and 6).Benefit: parallel processing allows the original and effected signals to be blended independently. See tips below.Tip: ,Example 1: crush the drums with heavy compression for tight, explosive impact, while mixing in the original to retain transients and dynamics.Tip: ,Example 2: add a touch of reverb to an entire mix to pull the group together in one space.Tip: ,Example 3: blend a dash of the original, unprocessed signal into the end of the chain for clarity.Tip: ,Example 4: mix in Dolby A in encode only to add some air (i.e. lead vocal on Journey’s, “Don’t Stop Believing” or for a more extreme example, Def Leppard Hysteria background vocals).Operation: Depress the parallel processing loop button to select the appropriate piece of outboard gear and engage flip buttons as needed.Tip: ,: Want to take advantage of parallel processing for insert1, not insert2? Simply reverse the order of the inserts with the8 | DANGEROUS MUSICflip1-2 switch. Now the insert1 button will blink slowly, indicating it’s activation in the parallel processing loop.6. PARALLEL PROCESSING LOOP BLEND CONTROL[parallel processing loop blend control pictured above]Feature: parallel processing amount controlled by a continuously variable potentiometer.Benefit: Dictates the amount of direct (non-effected) or processed (effected) signal that is sent to the end of buss b. See examples in item 4 above for application suggestions.7. POLARITY BUTTON[polarity button pictured above]Feature: Reverse the polarity 180 degrees.LIAISON | 9 Benefit: Something out of phase? Fix it. Need to dial something OUT of the mix? Add it here 180º out of phase and select the amount.Tip: ,As an effect: compress the mix and flip it out of phase to hear it pump in time with the track.8. PRESET, CLEAR & STORE BUTTONS[preset, clear and store buttons pictured above]Feature: store or clear up to four patches.Benefit: Have a favorite vocal processing chain? Master Buss sweetening path? Mastering outboard winning combination?Store all those proven success combinations and recall them instantly on demand.Operation: T o store a preset, select whatever combination of switches desired, press the store button (it will blink) and then depress the desired preset location.To clear a preset, simply select clear, then press store and the preset you’d like to clear.Tip: ,After a preset has been stored, changing any settings will cause the preset button to blink. Depressing the preset button will toggle between the stored preset (solid light) and modified preset (blinking light). Effectively doubling the memorylocations to 8. If desired, store the altered preset.Tip: ,Create presets as starting points. For example, when tracking, create your favorite vocal path, kick drum path etc...For the mix buss or mastering store presets for different musical styles (Hip Hop, Rock, Pop, Jazz, Gregorian Chant...)10 | DANGEROUS MUSICLiaison: Application Examples1. BLOCK DIAGRAMANALOG 8 CHANNEL DB25(TASCAM PROTOCOL)H = HOT C = COLD G = GROUNDPIN OUT WIRING FOR DB25 CONNECTORMONITOR OUT 1LEFT & RIGHT MONITOR OUT 2LEFT & RIGHT MONITOR OUT 3LEFT & RIGHT MONITOR OUT 4LEFT & RIGHTLIAISON |11STAND ALONE MASTERING EXAMPLEOUTBOARD GEAR INSERT 1OUTBOARD GEAR INSERT 2OUTBOARD GEAR INSERT 3OUTBOARD GEAR INSERT 4OUTBOARD GEAR INSERT 5OUTBOARD GEAR INSERT 612 |DANGEROUS MUSICMETERINGDANGEROUS MUSIC MONITOR DANGEROUS MUSIC MASTERLARGE FORMAT MASTERING EXAMPLESONTEC DRC400DANGEROUS MUSIC BAX EQSONTEC MES430BTOFT ATC2SONTEC DRC202MANLEY LABS VARIABLE MUGREAT RIVER Q2NVSONTEC M250EPLEXICON 960LLIAISON |13TRACKING & MIXING EXAMPLE AMONITOR STDANGEROUS MONITOR ST (OR MONITORING DEVICE)OUTBOARD GEAR INSERT 1OUTBOARD GEAR INSERT 2OUTBOARD GEAR INSERT 3OUTBOARD GEAR INSERT 4OUTBOARD GEAR INSERT 5OUTBOARD GEAR INSERT 614 |DANGEROUS MUSICSTAND ALONE MIXING EXAMPLEMIC PREAMPOUTBOARD GEAR INSERT 1OUTBOARD GEAR INSERT 2OUTBOARD GEAR INSERT 3OUTBOARD GEAR INSERT 4OUTBOARD GEAR INSERT 5OUTBOARD GEAR INSERT 6LIAISON |15TRACKING EXAMPLE ACOMPUTER INTERFACE (DAW/CONVERTER)*Utilizing input 4 on the MONITOR ST, allows the post processing level to be reduced, thus matching it to the original for true A/B comparisons.MONITOR STINPUT 1DANGEROUS MONITOR ST (OR MONITORING DEVICE)MIC PREAMPOUTBOARD GEAR INSERT 1OUTBOARD GEAR INSERT 2OUTBOARD GEAR INSERT 3OUTBOARD GEAR INSERT 4OUTBOARD GEAR INSERT 5OUTBOARD GEAR INSERT 616 |DANGEROUS MUSICTRACKING EXAMPLE B COMPUTER INTERFACE (DAW/CONVERTER)*Utilizing input 4 on the MONITOR ST, allows the post processing level to be reduced, thus matching it to the original for true A/B comparisons.MONITOR STINPUT 1DANGEROUS MONITOR ST (OR MONITORING DEVICE)MIC PREAMP 2MIC PREAMP 1OUTBOARD GEAR INSERT 1OUTBOARD GEAR INSERT 2OUTBOARD GEAR INSERT 3OUTBOARD GEAR INSERT 4OUTBOARD GEAR INSERT 5OUTBOARD GEAR INSERT 6LIAISON | 17 SPECIFICATIONS(for monitor and parallel buss sections)Frequency Response (Passive Section) +0, -0.1dB from 10Hz to 100kHzFrequency Response (Active Section) +0, -0.1dB 10Hz-100kHz+28dBu>MaximumlevelNoise floor < -94.5dBu 22Hz-22kHz bandwidth<0.002%THD+N0.003%<IMDCrosstalk rejection (Passive Section) > 124dBCrosstalk rejection (Active Section) > 113dBInput Impedance: 20K OhmsOutput Impedance: 20 OhmsReplacement Fuses: USA 500mA fast blow for 120VEurope 250mA fast blow for 240VNow it’s your turn to unleash the potential of the Liaison and tell us all the unusual applications you’ve utilized it in!Experiment well beyond the pages of this manual. The Liaison is a still audio river that runs deep.-The Dangerous Music Team18 | DANGEROUS MUSICWARRANTYFree 2 year extended warranty with online registration. Standard warranty: 90 days parts and labor, subject to inspection.Does not include damage incurred through shipping damage, abusive operation or modifications/attempted repair by unauthorized technicians.USA EUROPEDangerous Music Inc. Dangerous Music Europe231 Stevens Road Stieleichenweg 55Edmeston, NY 13335 50999 Cologne, GermanyFon: +49 2236 393731Email:***********************E-mail:**********************Dangerous Music, Inc. reserves the right to alter the software and design of their equipment.If after reading the manual more information for an application is needed, please contact us via email for the quickest response.•Factory contact for RA# must occur before shipping a unit to us for service.•Please keep the original cartons in case storage or transportation of units is required.•Always insure shipment as these damages are not covered by the warranty.•Thank you for actually reading the manual. Now go make some Dangerous Music!revision 5.0 marek April 2013。

L5-STUDIO用户手册（中文）说明书

简介在英国非常自豪的设计，工程和制造.狮心王（Lionheart）系列带来精品的单端或并联单端的甲类电子管音调提供给独具慧眼的演奏家，正是他们一直在寻找的一种独特的灵敏的具有英国特色电子管音调.L5-STUDIO是5W 甲类单端输出级渗出经典，温暖的电子管色调。

你驱动得越重，听起来就越好。

加上有足够的增益为现代的音调，它也有凶猛、侵略性的一面，使它适合任何风格的玩家. 完美的用于工作室和实践使用(使用5W或0.5W输出)，但同样适合在室内舞台上插入兰尼LT112、LT212或L412机柜。

这声音会让你大吃一惊。

利用在IRT-Studio中首次发布的T-USB技术，L5-STUDIO让你能够以一种前所未有的方式将电子管的色调和数字灵活的结合在一起，这对于那些在以前寻找复古美学，伟大的电子管音色与现代功能相结合的玩家来说是不可能的。

每一款狮心王 Lionheart产品都是由经验丰富的吉他手进行广泛的演奏测试，然后才发货给我们的客户。

只有当设备完成到我们完全满意时，它才被分配到自己的独特的建造编号，然后手工敲击在后面安装的铭牌上。

你的L5-STUDIO应该给你多年的无故障放大，然而，请花时间阅读本手册和熟悉自己的控制，因为它将允许你得到最好的放大器。

我们希望您喜欢使用您的L5-STUDIO，就像我们喜欢设计和制作它一样。

来自全体LANEY成员的最美好的祝福。

高输入:“HI”代表高增益。

这种输入是为低输出电平吉他的连接而设计的，使它非常适合单线圈和低增益双线圈型拾音器。

在这个输入中使用高增益拾取可能会使驱动前置放大器太硬，造成“模糊”的声音。

只能使用优质的吉他电缆. 低输入:“LO”代表低增益。

这种输入从高输入衰减了大约50%，是为高输出电平吉他设计的。

它对于从高增益双线圈型拾音器获得“紧密”而不是“糊状”的输出非常有用。

也使用这个输入为最干净的全范围的声音与扩展低端响应。

只能使用优质的吉他电缆.清音音量：设置清音通道的响度。

Audix i5 音响设备说明书

i5OVERVIEW:Designed, assembled and tested by Audix in the USA, the i5 is a dynamic instrument microphone used for stage, studio and broadcast applications. The i5 is able to handle sound pressure levels in excess of 140 dB without distortion and can be used to mic a wide variety of musical instruments, guitar and bass cabinets, vocals and speech.The i5 is characterized with a cardioid pickup pattern for isolation and feedback control and is equipped with a VLM™ (Very Low Mass) diaphragm for natural, accurate sound reproduction.The i5 is sturdy, compact and easy to position. With a wide frequency response of 50 Hz - 16 kHz, the i5 provides clear, accurate, natural sound reproduction without having to rely on EQ. Roadworthyconstruction includes a precision cast zinc alloy body, steel grill, black e-coat finish, laser etched model and serial number, Switchcraft® XLR connector and includes a heavy duty nylon mic clip.APPLICATIONS:Live stage, studioSnare, toms, percussionGuitar cabinets, electric instruments Bass cabinetsBrass, flute, woodwinds Acoustic instruments Vocals, speechDYNAMIC INSTRUMENT MICROPHONESUPPLIED ACCESSORIES:Heavy duty nylon mic clip (MC1)Carrying pouch (P1)OPTIONAL ACCESSORIES:DFLEX - All purpose percussion clamp DVICE - Spring loaded rim mount clamp DCLAMP - Tension rod mic clamp WS-i5 - External foam windscreen CBL-20 - 20’ XLR-XLR mic cableCBL-DR25 - 25’ right angle XLR mic cableCAB GRABBER - Tension held mic holder, clamps to guitar cabinetsDCLAMP DFLEX i5 on VOCALS CAB GRABBER i5 on SAXFEATURES:Clear and accurate sound reproduction Handles high SPL without distortionProvides exceptional gain before feedback Provides excellent isolation on stage Precision cast zinc alloy body and grill 5 year warrantyARCHITECTS AND ENGINEERS SPECIFICATIONS:The microphone shall be of the dynamic type operating on the moving coil principle and the polar pattern of the microphone shall be cardioid. The capsule shall be VLM Type B. The nominal out-put impedance shall be equal to 150 ohms at 1 kHz. The microphone shall have a sensitivity of 1.5 mV / Pa at 1 kHz and a sound pressure level ≥140 dB. The microphone body and grill shall be of die cast zinc alloy. The overall dimensions shall be 23 mm in diameter at the base, 37.5 mm at the top, and 141.5 mm in length. The microphone shall be the Audix i5. Call: 503-682-6933 Fax: 503-682-7114Audix Corporation 9400 SW Barber Street, Wilsonville, OR 97070Black E-coat Weight 183 g / 6.5 ounces Length 141.5 mm / 5.6 inchesOPERATION:The i5 is a low impedance microphone and should be plugged into a “mic level” input on your console, mixer, or recording device. Please note that your microphone does not require phantom power and will not be effected in any way by phantom power should it be running simultaneously when the microphone is in operation. Avoid plugging or unplugging the microphone from the PA system unless the channel is muted or the volume of the system turned down. Failure to do so may result in a loud “popping” noise which could seriously damage the speakers in the PA system.FREQUENCY / POLARS:To register your microphone, please visit SERVICE AND WARRANTY:This microphone is under warranty for a period of 5 years from any and all manufacturing defects. Should your microphone fail in any way, please con-tact the Audix Service department at 503-682-6933. A Return Authorization number is required before returning any products .CARE AND MAINTENANCE:The i5 is manufactured to exacting specs with roadworthy construction. However, the capsule is highly sensitive and should be handled with care. Avoid extreme temperatures and be sure to store your microphone in the pouch provided when not in use. Moisture of any kind can adversely affect the sound and performance of your microphone.***All specifications subject to change without notice.USER TIPS:The i5 has a tight (cardioid) pick-up pattern in order to help eliminate sound from other instruments on stage from “bleeding” into the microphone. For this reason, it the i5 is exceptionally effective for drums and percussion in be able to capture the sound of each component of the kit.Snare: Start with the i5 about 2 inches above the head and aimed towards the center of the drum. For more “rim” effect, the i5 can be aimed less towards the center and more towards the rim. It is not uncommon for engineers to place the mic practically right on the drum head to achieve a very tight, cracking sensation.Toms, congas: Similar to snare. For tighter sound and more attack place, the mic closer to the head. For more decay and overtones pull the mic further away from the head.Guitar cabs: The i5 can be placed within 1-2 inches of the grill cover at a 90 degree angle pointing directly into the speaker. If the mic is placed closer to the edge of the speaker, you will minimize the higher frequencies and get a warmer, fatter tone. To capture more highs, move the mic closer towards the center of the speaker, but avoid the middle of the speaker.Horns: Place the mic within 1-3 inches of the bell, especially when on a stage with drums, bass, guitar, etc. For jazz, classical, and music where the stage volume is lower, the mic can be placed further away for a richer, fuller, sound.Allow a distance of 2-3 feet between microphones to avoid phase cancellation issues. *Further miking techniques may be found on our website at DIMENSIONS (mm):。

蒂尔斯菲利奥5.1.2声音条说明书

Philips Fidelio Soundbar 5.1.2 with wireless subwoofer808W max. Wireless subwoofer Dolby Atmos. IMAX Enhanced DTS Play-Fi compatible Connects with voice assistantsB95Stunning sound for every drama. Or song.Experience your movies and music like never before. This Fidelio soundbar with wireless subwoofer brings new levels of detail to every scene and soundtrack. Plus, it’s easy to make this soundbar part of your multi-room audio setup.Give your entertainment the sound it deserves•Philips Fidelio. Stunning sound with rich, deep bass•Dolby Atmos and Dolby Surround. Heighten the excitement•5.1.2 channels. 808 W max. 410 W RMS (1%THD)•Dedicated center channel for crystal-clear dialogueNew levels of detail for movies and music•IMAX Enhanced. Immersive, heart-pounding audio•Play-Fi. From multi-room audio to surround sound•HDMI eARC. Enjoy Dolby Atmos and DTS:X audio formats•Spotify Connect. Bluetooth. Apple AirPlay 2. Optical-inA seamless experience•Effortless. From movie night to music•Easily connect to your favorite sources•4K pass-through. Dual HDMI inputs•Connects to your favorite voice assistant-enabled devicesEffortless connectionWith a Philips Fidelio soundbar, nothing comes between you and the moment. Whether it's the seamless way you can switch between all your favorite sources or connect to speakers in other rooms. Or the low-profile design that allows easy placement. Simply unpack, connect and enjoy.Stunning sound. Rich deep bassThis soundbar creates a breathtaking sound experience. 5.1.2 channels fill any room with rich, deep soundtracks, thunderous effects, and sparkling dialogue. The wireless subwoofer adds impact to every explosion and every beat.Dolby Atmos and Dolby SurroundGet the full Dolby Atmos experience from this soundbar's up-firing speakers. Your favoritemovies and shows will sound breathtakingly realistic as sound flows above and around you. If you're watching non-Atmos content, the soundbar switches to Dolby Surround.IMAX EnhancedThis powerful soundbar is certified IMAX Enhanced, which means it's capable ofdelivering an audio experience that truly takes you to another world. The 240 W subwoofer looks great freestanding, and the soundbar itself has a low profile for simple placement.Play-Fi. Multi-room and moreEasy Play-Fi compatibility will make thissoundbar the star of your multi-room set-up. You can sync Play-Fi-compatible speakers for multi-room audio, or even create a true surround-sound system—all via the Philips Sound app, or the Play-Fi app.All your favorite sourcesFor music, Spotify Connect lets you stream Spotify's best-quality signal over Wi-Fi. You can also stream hi-res playlists from your mobile device via Apple AirPlay 2 or Bluetooth. For movies and gaming, 4K pass-through lets youconnect 4K HDR video sources with no loss of resolution.HDMI eARCLose nothing from the mix when you're immersed in the drama. This soundbar is compatible with HDMI eARC, a high-speed connection that lets you experience the full effect of advanced audio formats like Dolby Atmos and DTS:X. The soundbar also has dual HDMI inputs.Connect with voice assistantsAsk Alexa-enabled devices, or any speaker that works with the Google Assistant, to play music through the soundbar. Prefer Siri? Thissoundbar works with Apple AirPlay 2, so you can ask her too. Turn the music up. Skip tracks.All completely hands-free.Issue date 2022-11-02Version: 8.0.112 NC: 8670 001 73446UPC: 8 40063 20167 5© 2022 Koninklijke Philips N.V.All Rights reserved.Specifications are subject to change without notice. Trademarks are the property of Koninklijke Philips N.V. or their respective owners.Loudspeakers•Number of sound channels: 5.1.2•Center drivers: 2 full range (L+R)•Front drivers: 4 full range (L+R), 2 tweeters (L+R)•Up-firing drivers: 2 full range (L+R)•Side-firing drivers: 2 tweeters (L+R)•Soundbar freq range: 150 - 20k Hz•Subwoofer type: Active, Wireless subwoofer •External subwoofer enclosure: Bass reflex •Number of woofers: 1•Woofer diameter: 8"•Subwoofer freq range: 35 - 150 Hz •Subwoofer impedance: 3 ohmConnectivity•HDMI IN x 2•HDMI Out (eARC/ARC) x 1•EasyLink (HDMI-CEC)•HDMI content protection: HDCP 1.4/2.3•Optical input x 1•Aux in: 3.5mm•Bluetooth: Receiver •Bluetooth version: 4.2•DLNA Standard:No•Multiple speaker connection•Multiple speaker technology: DTS Play-Fi Multi-room•Smart Home: Built-in Chromecast, SpotifyConnect, Works with Alexa, Works with Apple AirPlay 2, Works with OK Google•WiFi: 2.4GHz/5GHz, IEEE 802.11 a/b/g/n/ac •Wireless speaker connection: SubwooferSound•Sound System:Dolby Atmos, DTS:X, IMAX Enhanced•Sound Enhancement: Movie, Music, Voice, Sport, Custom•Speaker system output power: 808W MAX / 410W RMS (1%THD)•Total harmonic distortion: 1%Video•Video Enhancement: 4K Video Pass-through, Dolby Vision, HDR10, HDR10+, HLG •3D pass-throughSupported Audio Formats•HDMI IN1/IN2: Dolby Atmos, Dolby Digital,Dolby Digital plus, Dolby TrueHD, Dolby Digital EX, DTS, DTS 96/24, DTS Digital Surround (5.1-ch), DTS:X, DTS-ES, DTS-HD High Resolution Audio, DTS-HD Master Audio•HDMI ARC/eARC: Dolby Atmos, Dolby Digital, Dolby Digital EX, Dolby Digital plus, DolbyTrueHD, DTS, DTS 96/24, DTS Digital Surround (5.1-ch), DTS:X, DTS-ES, DTS-HD High Resolution Audio, DTS-HD Master Audio •Optical: Dolby Digital, LPCM 2ch •Bluetooth:SBC•Play-Fi: AIFF, FLAC, MP3, WAV, MPEG4 AAC, MPEG4 M4ACompatibility •Smartphone/tablet APP control Convenience•Night mode:No •Remote ControlDesign•Color: Black •Wall mountablePower•Main unit power supply: 100-240V AC, 50/60 Hz •Main unit standby power: < 0.5 W•Subwoofer Power supply: 100-120V AC, 60 Hz •Subwoofer standby power: < 0.5 W •Auto standbyAccessories•Included accessories: Remote Control (with battery), Power cord, Quick start guide, World Wide Warranty leafletDimensions•Main Unit (W x H x D): 1049 x 55.6 x 120 mm •Main Unit Weight: 4.07 kg•Subwoofer (W x H x D): 230 x 400 x 407 mm •Subwoofer Weight: 9.6 kgPackaging dimensions•UPC: 8 40063 20167 5•Packaging dimensions (W x H x D): 115 x 45.8 x 40.5 cm•Packaging dimensions (W x H x D): 45.3 x 18.0 x 15.9 inch•Gross weight: 41.446 lb •Gross weight: 18.8 kg •Nett weight: 13.87 kg •Nett weight: 30.578 lb•Number of products included: 1•Packaging type: Box •Tare weight: 4.93 kg •Tare weight: 10.869 lbOuter Carton•GTIN: 1 08 40063 20167 2•Number of consumer packagings: 2•Outer carton (L x W x H): 45.9 x 21.1 x 18.9 inch •Outer carton (L x W x H): 116.5 x 53.5 x 48 cm •Gross weight: 88.845 lb •Gross weight: 40.3 kg •Nett weight: 27.74 kg •Nett weight: 61.156 lb •Tare weight: 27.690 lb •Tare weight: 12.56 kg。

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User Manual
Version 1.0
Revision Date: 08/12/2013 Product name: I-5™ 2X High-Fidelity Master Mix
Cat #: I5HM-200, I5HM-5000, I5HM-OEM
Description:
I-5™ High-Fidelity DNA Polymerases is an ultra-high fidelity and high processivity enzyme. It produces the most accurate copies of DNA, and performs at an ultra-high rate. I-5™ is perfect for applications like cloning in vitro amplified material for protein expression, SNP analysis by sequencing, and high-specificity PCR.
Convenient and Usage:
I-5™ 2X High-Fidelity Master Mix has a buffer system that is designed to offer maximum performance of the I-5™ High-Fidelity DNA Polymerase. It offers the highest fidelity and incredibly robust performance. The convenient master mix allows reaction to be set up in room temperature and only requires the additions of primers and DNA template for the amplification.
Application:
- High-specificity PCR amplification
- High-Throughput PCR
- Various Cloning technologies
- Difficult Amplification
Recommended Storage Conditions:
-20°C.
Instructions:
25 μl Reaction 50 μl Reaction Final Concentration
I-5TM 2X High-Fidelity Master Mix 12.5 µl 25 µl 1X (see notes)
10 μM Primer A 1 µl 2 µl 400 µM
10 μM Primer B 1 µl 2 µl 400 µM
Template DNA as required as required see notes
Water ddH2O up to 25 μl up to 50 μl
Notes:
Please do pre experimental verification in other reaction volume.
Thermocycling Conditions
Step Temperature Time
Initial Denaturation 98℃ 2 minutes
Denaturation 98℃10 seconds 25-35 cycles
Annealing 45℃-68℃10-15 seconds
Extension 72℃5-15 seconds / kb
Final Extension 72℃1-5 minutes
4℃Hold
Notes:
Genomic DNA 50-250 ng
Plasmid DNA 1 pg-10 ng
Viral DNA 1 pg-10 ng
The time of extension depends on the type of PCR instrument, but not less than10 seconds.
Mg2+
The final concentration of Mg2+in 1X I-5™ PCR Master Mix is 2 mM
Add additional Mg2+as needed in 0.5 mM increments
Suggested final Mg2+concentration ranges from 2 mM to 4 mM
PCR Product / Cloning
The 2X I-5™ PCR Master Mix results in PCR product with blunt ends
Blunt-end cloning is recommended after PCR with this product
For T/A-cloning, the PCR product should be purified before A-addition。