pAcGFP1-F哺乳动物表达载体说明

pAcGFP1-F编号 载体名称北京华越洋生物VECT6054 pAcGFP1-­‐F pAcGFP1-­‐F载体基本信息载体名称: pAcGFP1-F质粒类型: 哺乳动物细胞表达载体；荧光报告载体高拷贝/低拷贝: 高拷贝克隆方法: 限制性内切酶，多克隆位点启动子: CMV IE载体大小: 4806 bp5' 测序引物及序列: --3' 测序引物及序列: --载体标签: AcGFP1 (N-端）载体抗性: 卡那霉素筛选标记: 新霉素（Neomycin）克隆菌株: DH5α, HB101宿主细胞（系）: 常规细胞系（293、CV-1、CHO等）备注: 哺乳动物载体pAcGFP1-F组成型表达Ras-AcGFP融合蛋白；融合蛋白定位于细胞内膜上，荧光标记细胞膜。

稳定性: 稳表达或瞬表达组成型/诱导型: 组成型病毒/非病毒: 非病毒pAcGFP1-­‐F载体质粒图谱和多克隆位点信息pAcGFP1-F载体描述Ras (1, 2) fused to the C-terminus of AcGFP1. Post-translation of this farnesylation signal targets AcGFP1-F to the inner leaflet of the plasma membrane. AcGFP1 is derived from Aequorea coerulescens. When AcGFP1-F is expressed in mammalian cell cultures, green fluorescent cells can be detected by either fluorescence microscopy or flow cytometry 12–16 hr after transfection (excitation maximum = 475 nm; emission maximum = 505 nm, respectively). The AcGFP1 coding sequence is human-codon-optimized for increased translation efficiency in mammalian cells (3). SV40 polyadenylation signals downstream of the AcGFP1-F gene direct proper processing of the 3’ end of the AcGFP1-F mRNA. The vector backbone also contains an SV40 origin for replication in mammalian cells expressing the SV40 T-antigen. A neomycin resistance cassette (Neor), consisting of the SV40 early promoter, the neomycin/kanamycin resistance gene of Tn5, and polyadenylation signals from the herpes simplex virus thymidine kinase (HSV TK) gene, allows stably transfected eukaryotic cells to be selected using G418. A bacterial promoter upstream of this cassette expresses kanamycin resistance in E. coli. The pAcGFP1-F backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production.pAcGFP1-F is designed for use as a plasma membrane marker, as well as a cotransfection marker. Because it remains attached to the plasma membrane, it can be detected by fluorescence microscopy in permeabilized cells after ethanol fixation (4). The vector can be transfected into mammalian cells using any standard transfection method. If required, stable transformants can be selected using G418 (5). Propagation in E. coliSuitable host strains: DH5a, HB101, and other general purpose strains. Single-stranded DNA productionrequires a host containing an F plasmid such as JM109 or XL1-Blue.Selectable marker: plasmid confers resistance to kanamycin (50 μg/ml) in E. coli hosts.E. coli replication origin: pUC; copy number: highReferences1. Aronheim, A., et al. (1994) Cell 78:949–961.2. Hancock, J. F., et al. (1991) EMBO J. 10:4033–4039.3. Haas, J., et al. (1996) Curr. Biol. 6:315–324.4. Jiang, W. & Hunter, T. (1998) BioTechniques 24:348–354.5. Gorman, C. (1985) In DNA Cloning: A Practical Approach, Vol. II, Ed. Glover, D. M. (IRL Press, Oxford, UK) pp. 143–190.Note: The attached sequence file has been compiled from information in the sequence databases, published literature, and other sources,together with partial sequences obtained by Clontech. This vector has not been completely sequenced.pAcGFP1-F载体序列ORIGIN1 TAGTTATTAA TAGTAATCAA TTACGGGGTC ATTAGTTCAT AGCCCATATA TGGAGTTCCG 61 CGTTACATAA CTTACGGTAA ATGGCCCGCC TGGCTGACCG CCCAACGACC CCCGCCCATT 121 GACGTCAATA ATGACGTATG TTCCCATAGT AACGCCAATA GGGACTTTCC ATTGACGTCA 181 ATGGGTGGAG TATTTACGGT AAACTGCCCA CTTGGCAGTA CATCAAGTGT ATCATATGCC 241 AAGTACGCCC CCTATTGACG TCAATGACGG TAAATGGCCC GCCTGGCATT ATGCCCAGTA 301 CATGACCTTA TGGGACTTTC CTACTTGGCA GTACATCTAC GTATTAGTCA TCGCTATTAC 361 CATGGTGATG CGGTTTTGGC AGTACATCAA TGGGCGTGGA TAGCGGTTTG ACTCACGGGG 421 ATTTCCAAGT CTCCACCCCA TTGACGTCAA TGGGAGTTTG TTTTGGCACC AAAATCAACG 481 GGACTTTCCA AAATGTCGTA ACAACTCCGC CCCATTGACG CAAATGGGCG GTAGGCGTGT 541 ACGGTGGGAG GTCTATATAA GCAGAGCTGG TTTAGTGAAC CGTCAGATCC GCTAGCGCTA 601 CCGGTCGCCA CCATGGTGAG CAAGGGCGCC GAGCTGTTCA CCGGCATCGT GCCCATCCTG 661 ATCGAGCTGA ATGGCGATGT GAATGGCCAC AAGTTCAGCG TGAGCGGCGA GGGCGAGGGC 721 GATGCCACCT ACGGCAAGCT GACCCTGAAG TTCATCTGCA CCACCGGCAA GCTGCCTGTG 781 CCCTGGCCCA CCCTGGTGAC CACCCTGAGC TACGGCGTGC AGTGCTTCTC ACGCTACCCC 841 GATCACATGA AGCAGCACGA CTTCTTCAAG AGCGCCATGC CTGAGGGCTA CATCCAGGAG901 CGCACCATCT TCTTCGAGGA TGACGGCAAC TACAAGTCGC GCGCCGAGGT GAAGTTCGAG 961 GGCGATACCC TGGTGAATCG CATCGAGCTG ACCGGCACCG ATTTCAAGGA GGATGGCAAC 1021 ATCCTGGGCA ATAAGATGGA GTACAACTAC AACGCCCACA ATGTGTACAT CATGACCGAC 1081 AAGGCCAAGA ATGGCATCAA GGTGAACTTC AAGATCCGCC ACAACATCGA GGATGGCAGC 1141 GTGCAGCTGG CCGACCACTA CCAGCAGAAT ACCCCCATCG GCGATGGCCC TGTGCTGCTG 1201 CCCGATAACC ACTACCTGTC CACCCAGAGC GCCCTGTCCA AGGACCCCAA CGAGAAGCGC 1261 GATCACATGA TCTACTTCGG CTTCGTGACC GCCGCCGCCA TCACCCACGG CATGGATGAG 1321 CTGTACAAGT CCGGACTCAG ATCTAAGCTG AACCCTCCTG ATGAGAGTGG CCCCGGCTGC 1381 ATGAGCTGCA AGTGTGTGCT CTCCTGAGGA TCCAGATCTC GAGCTCAAGC TTCGAATTCT 1441 GCAGTCGACG GTACCGCGGG CCCGGGATCC ACCGGATCTA GATAACTGAT CATAATCAGC 1501 CATACCACAT TTGTAGAGGT TTTACTTGCT TTAAAAAACC TCCCACACCT CCCCCTGAAC 1561 CTGAAACATA AAATGAATGC AATTGTTGTT GTTAACTTGT TTATTGCAGC TTATAATGGT 1621 TACAAATAAA GCAATAGCAT CACAAATTTC ACAAATAAAG CATTTTTTTC ACTGCATTCT 1681 AGTTGTGGTT TGTCCAAACT CATCAATGTA TCTTAACGCG TAAATTGTAA GCGTTAATAT 1741 TTTGTTAAAA TTCGCGTTAA ATTTTTGTTA AATCAGCTCA TTTTTTAACC AATAGGCCGA 1801 AATCGGCAAA ATCCCTTATA AATCAAAAGA ATAGACCGAG ATAGGGTTGA GTGTTGTTCC 1861 AGTTTGGAAC AAGAGTCCAC TATTAAAGAA CGTGGACTCC AACGTCAAAG GGCGAAAAAC 1921 CGTCTATCAG GGCGATGGCC CACTACGTGA ACCATCACCC TAATCAAGTT TTTTGGGGTC 1981 GAGGTGCCGT AAAGCACTAA ATCGGAACCC TAAAGGGAGC CCCCGATTTA GAGCTTGACG 2041 GGGAAAGCCG GCGAACGTGG CGAGAAAGGA AGGGAAGAAA GCGAAAGGAG CGGGCGCTAG 2101 GGCGCTGGCA AGTGTAGCGG TCACGCTGCG CGTAACCACC ACACCCGCCG CGCTTAATGC 2161 GCCGCTACAG GGCGCGTCAG GTGGCACTTT TCGGGGAAAT GTGCGCGGAA CCCCTATTTG 2221 TTTATTTTTC TAAATACATT CAAATATGTA TCCGCTCATG AGACAATAAC CCTGATAAAT 2281 GCTTCAATAA TATTGAAAAA GGAAGAGTCC TGAGGCGGAA AGAACCAGCT GTGGAATGTG 2341 TGTCAGTTAG GGTGTGGAAA GTCCCCAGGC TCCCCAGCAG GCAGAAGTAT GCAAAGCATG 2401 CATCTCAATT AGTCAGCAAC CAGGTGTGGA AAGTCCCCAG GCTCCCCAGC AGGCAGAAGT 2461 ATGCAAAGCA TGCATCTCAA TTAGTCAGCA ACCATAGTCC CGCCCCTAAC TCCGCCCATC 2521 CCGCCCCTAA CTCCGCCCAG TTCCGCCCAT TCTCCGCCCC ATGGCTGACT AATTTTTTTT 2581 ATTTATGCAG AGGCCGAGGC CGCCTCGGCC TCTGAGCTAT TCCAGAAGTA GTGAGGAGGC 2641 TTTTTTGGAG GCCTAGGCTT TTGCAAAGAT CGATCAAGAG ACAGGATGAG GATCGTTTCG 2701 CATGATTGAA CAAGATGGAT TGCACGCAGG TTCTCCGGCC GCTTGGGTGG AGAGGCTATT 2761 CGGCTATGAC TGGGCACAAC AGACAATCGG CTGCTCTGAT GCCGCCGTGT TCCGGCTGTC 2821 AGCGCAGGGG CGCCCGGTTC TTTTTGTCAA GACCGACCTG TCCGGTGCCC TGAATGAACT 2881 GCAAGACGAG GCAGCGCGGC TATCGTGGCT GGCCACGACG GGCGTTCCTT GCGCAGCTGT 2941 GCTCGACGTT GTCACTGAAG CGGGAAGGGA CTGGCTGCTA TTGGGCGAAG TGCCGGGGCA 3001 GGATCTCCTG TCATCTCACC TTGCTCCTGC CGAGAAAGTA TCCATCATGG CTGATGCAAT 3061 GCGGCGGCTG CATACGCTTG ATCCGGCTAC CTGCCCATTC GACCACCAAG CGAAACATCG 3121 CATCGAGCGA GCACGTACTC GGATGGAAGC CGGTCTTGTC GATCAGGATG ATCTGGACGA 3181 AGAGCATCAG GGGCTCGCGC CAGCCGAACT GTTCGCCAGG CTCAAGGCGA GCATGCCCGA 3241 CGGCGAGGAT CTCGTCGTGA CCCATGGCGA TGCCTGCTTG CCGAATATCA TGGTGGAAAA 3301 TGGCCGCTTT TCTGGATTCA TCGACTGTGG CCGGCTGGGT GTGGCGGACC GCTATCAGGA 3361 CATAGCGTTG GCTACCCGTG ATATTGCTGA AGAGCTTGGC GGCGAATGGG CTGACCGCTT 3421 CCTCGTGCTT TACGGTATCG CCGCTCCCGA TTCGCAGCGC ATCGCCTTCT ATCGCCTTCT 3481 TGACGAGTTC TTCTGAGCGG GACTCTGGGG TTCGAAATGA CCGACCAAGC GACGCCCAAC3541 CTGCCATCAC GAGATTTCGA TTCCACCGCC GCCTTCTATG AAAGGTTGGG CTTCGGAATC3601 GTTTTCCGGG ACGCCGGCTG GATGATCCTC CAGCGCGGGG ATCTCATGCT GGAGTTCTTC3661 GCCCACCCTA GGGGGAGGCT AACTGAAACA CGGAAGGAGA CAATACCGGA AGGAACCCGC3721 GCTATGACGG CAATAAAAAG ACAGAATAAA ACGCACGGTG TTGGGTCGTT TGTTCATAAA3781 CGCGGGGTTC GGTCCCAGGG CTGGCACTCT GTCGATACCC CACCGAGACC CCATTGGGGC3841 CAATACGCCC GCGTTTCTTC CTTTTCCCCA CCCCACCCCC CAAGTTCGGG TGAAGGCCCA3901 GGGCTCGCAG CCAACGTCGG GGCGGCAGGC CCTGCCATAG CCTCAGGTTA CTCATATATA3961 CTTTAGATTG ATTTAAAACT TCATTTTTAA TTTAAAAGGA TCTAGGTGAA GATCCTTTTT4021 GATAATCTCA TGACCAAAAT CCCTTAACGT GAGTTTTCGT TCCACTGAGC GTCAGACCCC4081 GTAGAAAAGA TCAAAGGATC TTCTTGAGAT CCTTTTTTTC TGCGCGTAAT CTGCTGCTTG4141 CAAACAAAAA AACCACCGCT ACCAGCGGTG GTTTGTTTGC CGGATCAAGA GCTACCAACT4201 CTTTTTCCGA AGGTAACTGG CTTCAGCAGA GCGCAGATAC CAAATACTGT TCTTCTAGTG4261 TAGCCGTAGT TAGGCCACCA CTTCAAGAAC TCTGTAGCAC CGCCTACATA CCTCGCTCTG4321 CTAATCCTGT TACCAGTGGC TGCTGCCAGT GGCGATAAGT CGTGTCTTAC CGGGTTGGAC4381 TCAAGACGAT AGTTACCGGA TAAGGCGCAG CGGTCGGGCT GAACGGGGGG TTCGTGCACA4441 CAGCCCAGCT TGGAGCGAAC GACCTACACC GAACTGAGAT ACCTACAGCG TGAGCTATGA4501 GAAAGCGCCA CGCTTCCCGA AGGGAGAAAG GCGGACAGGT ATCCGGTAAG CGGCAGGGTC4561 GGAACAGGAG AGCGCACGAG GGAGCTTCCA GGGGGAAACG CCTGGTATCT TTATAGTCCT4621 GTCGGGTTTC GCCACCTCTG ACTTGAGCGT CGATTTTTGT GATGCTCGTC AGGGGGGCGG4681 AGCCTATGGA AAAACGCCAG CAACGCGGCC TTTTTACGGT TCCTGGCCTT TTGCTGGCCT4741 TTTGCTCACA TGTTCTTTCC TGCGTTATCC CCTGATTCTG TGGATAACCG TATTACCGCC4801 ATGCAT//其他哺乳动物表达载体：pCHO1.0 pBApo-CMV-Pur pOPRSVIpcDNA3.1/His C pcDNA5/FRT/V5-His-TOPO pREP4pcDNA3.1/His B pcDNA5/FRT/TO-TOPO pDual-GCpcDNA3.1/His A pcDNA5/TO pBK-RSVpIRESpuro3 pcDNA5/FRT/TO pBK-CMVpIRES2-EGFP pcDNA5/FRT pBI-CMV4pTT5 pFLAG-CMV2 pcDNA4/TO/Myc-His/LacZ pNFkB-DD-tdTomato pcDNA3.1/CT-GFP-TOPO pOPI3CATpBI-CMV5 pcDNA3.1/NT-GFP-TOPO pGene/V5-His B pSEAP2-Basic pOptiVEC-TOPO pSwitchpSEAP2-Control pCMV-MEKK1 pCMVLacIpBI-CMV3 pCMV-MEK1 pVgRxRpBI-CMV2 pCMV-PKA pINDpBI-CMV1 pcDNA6.2/nTC-Tag-DEST pTRE3G-LucpNFκB-MetLuc2-Reporter pcDNA6.2/cTC-Tag-DEST pTRE3GpCRE-MetLuc2-Reporter pcDNA3.2/V5/GW/D-TOPO pTRE2-hygropAcGFP1-Actin pcDNA6.2/V5/GW/D-TOPO pTRE-TightpAcGFP1-N In-Fusion Ready pcDNA6.2/nGeneBLAzer-GW/D-TOPO pTK-hygpAcGFP1-C3 pcDNA6.2/C-YFP-DEST pTet-OnpAcGFP1-C pcDNA6.2/cGeneBLAzer-DEST pTet-OffpAcGFP1-p53 pcDNA6/V5-His A pTet on advanced pAcGFP1-Mito pcDNA6/V5-His B pRevTREpAcGFP1-Mem pcDNA6/V5-His C pRevTet-On pAcGFP1-Lam pcDNA6/myc-His C pRevTet-Off pAcGFP1-Golgi pcDNA6/myc-His A pCMV-Tet3G pAcGFP1-F pcDNA6/myc-His B pTRE2pAcGFP1-Hyg-C1 pcDNA6.2/nGeneBLAzer-DEST pBD-NF-κB pAsRed2-N1 pcDNA4/HisMax-TOPO pCMV-AD ptdTomato-N1 pcDNA6.2/nLumio-DEST pCMV-BDpCMV-tdTomato pcDNA6.2/cLumio-DEST pBIND-Id Control pCRE-DD-tdTomato pcDNA4/myc-His C pBINDpCMV-DsRed-Express2 pcDNA4/HisMax C pG5 luciferasepEF1α-tdTomato pcDNA4/HisMax A pACT-MyoDpCRE-hrGFP c-Flag pcDNA3 pACTptdTomato-C1 pcDNA4/HisMax B pCMV-SPORT6 pAsRed2-C1 pcDNA4/myc-His B pGL4.13pGL3-Promoter pcDNA4/myc-His A pGL4.19pGL3 basic pcDNA4/His C pGL4.26pAcGFP1-C2 pcDNA4/His B pGL4.20pAcGFP1-C1 pcDNA4/His A pGL4.29pAcGFP1-N3 pcDNA6/TR pGL4.30pAcGFP1-N2 pcDNA4/TO/Myc-His A pGL4.27pAcGFP1-N1 pcDNA4/TO pGL4.75pAcGFP1-C In-Fusion Ready pcDNA4/TO/Myc-His B pGL4.10pCRE-DD-AmCyan1 pcDNA4/TO/Myc-His C pGRN145pNFkB-DD-AmCyan1 pcDNA3.3-TOPO pSecTag2 A pDsRed2-Bid pBudCE4.1 pEBVHis B pDsRED2-Mito pFLAG-CMV-4 pEBVHis ApDD-AmCyan1 Reporter pFLAG-CMV-3 pCMV-Tag 3C pAmCyan1-N1 pFLAG-CMV-2 pCMV-Tag 3A pAmCyan1-C1 pFLAG-CMV-5a pCMV-Tag 3BpEF1α-IRES-DsRed-Express2 p3XFLAG-CMV-9 pCMV-Tag 5CpEF1α-DsRed-Monomer-N1 p3xFLAG-CMV-10 pCMV-Tag 5A pDsRED-Monomer-N1 p3XFLAG-CMV-8 pCMV-Tag 4A pDsRed-Express-N1 p3XFLAG-CMV-7.1 pCMV-Tag 5Bp3XFLAG-CMV-7 pDsRed-Monomer-N In-Fusion Ready pCMV-Tag 4B pDsRed-Express-C1 p3XFLAG-CMV-13 pCMV-Tag 2C pIRES2-ZsGreen1 p3XFLAG-CMV-14 pCMV-Tag 2B pDsRed-Express2-C1 plRES2-ZsGreen1 pCMV-Tag 2A pDsRed-Express2-N1 pBApo-EF1α-pur pCMV-LacZpEF1α-DsRed-Express2 pBApo-EF1α-neo pCMV-MycpIRES2-DsRed-Express pBApo-CMV pEF1α-IRES-AcGFP1 pIRES2-DsRed-Express2 pBApo-CMV-neo pEF1α-IRES-ZsGreen1pIRES-hrGFP-1a pIRES-EGFP pEF1α-AcGFP1-N1 pIRESneo2 pIRESneo3 pIRES2-DsRed2 pIRESneo pDsRed-Monomer pIRES2-AcGFP1 pIREShyg3 pIRES。