pLVX-DsRed-Monomer-N1慢病毒载体使用说明

pLVX-DsRed-Monomer-N1

pLVX-DsRed-Monomer-N1载体基本信息：

pLVX-DsRed-Monomer-N1载体质粒图谱和多克隆位点信息：

载体名称: pLVX-DsRed-Monomer-N1

质粒类型: 慢病毒表达载体；荧光报告载体

高拷贝/低拷贝: 高拷贝

启动子: CMV

克隆方法: 多克隆位点，限制性内切酶

载体大小: 8751 bp

5' 测序引物及序列: CMV-F: CGCAAATGGGCGGTAGGCGTG (Invitrogen)

3' 测序引物及序列: --

载体标签: DsRed-Monomer（C-端）

载体抗性: 氨苄青霉素

筛选标记: 嘌呤霉素(Puromycin)

克隆菌株: Stbl3

宿主细胞（系）: 常规细胞系，293、CV-1、CHO等

备注:

pLVX-DsRed-Monomer-N1载体表达C端DsRed-Monomer

融合蛋白；

DsRed-Monomer是DsRed的单体突变体，与DsRed相比，

编码序列经过了人工优化，适用于在哺乳动物细胞中的

高水平表达；

CMV启动子是过表达启动子。

稳定性: 稳表达

组成型: 组成型

病毒/非病毒: 慢病毒

pLVX-DsRed-Monomer-N1载体简介：

Description

pLVX-DsRed-Monomer-N1 is an HIV-1-based, lentiviral expression vector that allows you to express your gene of interest fused to DsRed-Monomer, a monomeric mutant of the Discosoma sp. red fluorescent protein. Genes cloned into the multiple cloning site (MCS),located upstream of the DsRed-Monomer coding sequence, are expressed as N-terminal fusions of the DsRed-Monomer protein. Expression of the fusion protein is driven by the constitutively active human cytomegalovirus immediate early promoter (PCMV IE) located just upstream of the MCS. Lentiviral particles derived from the vector allow the expression of DsRed-Monomer fusion proteins in virtually any cell type, including primary cells. The unmodified vector expresses

DsRed-Monomer, and may be used to produce marker virus to

optimize infection protocols.

pLVX-DsRed-Monomer-N1 contains all of the viral processing elements necessary for the production of replication-incompetent lentivirus, as well as elements to improve viral titer, transgene expression, and overall vector function. The woodchuck hepatitis virus posttranscriptional regulatory element (WPRE) promotes RNA processing events and nhances nuclear export of viral and transgene RNA (1), leading to increased viral titers from packaging cells, and enhanced expression of your gene of interest in target cells. In addition, the vector includes a Rev-response element (RRE), which further increases viral titers by enhancing the transport of unspliced viral RNA out of the nucleus (2). Finally, pLVX-DsRed-Monomer-N1 also contains a central polypurine tract (cPPT) element that increases nuclear importation of the viral genome during target cell infection, resulting in improved vector integration and

more efficient transduction (3).

In addition to lentiviral elements, pLVX-DsRed-Monomer-N1 contains a puromycin resistance gene (Puror) under the control of the murine phosphoglycerate kinase (PGK) promoter (PPGK) for the selection of stable transductants. The vector also contains a pUC origin of replication and an E. coli ampicillin resistance gene (Ampr) for propagation and selection in bacteria.

Use

pLVX-DsRed-Monomer-N1 constitutively expresses your gene of interest from PCMV IE when transduced into target cells. Before the vector can be transduced into cells, however, it must be transfected into 293T packaging cells with our Lenti-X™ HTX Packaging System (Cat. Nos. 631247 and 631249). This packaging system allows you to safely produce high titer, infectious, replication-incompetent, VSV-G pseudotyped lentiviral particles that can infect a wide range of cell types, including non-dividing and primary cells (4).

pLVX-DsRed-Monomer-N1载体序列：

ORIGIN

1 TGGAAGGGCT AATTCACTCC CAAAGAAGAC AAGATATCCT TGATCTGTGG ATCTACCACA

61 CACAAGGCTA CTTCCCTGAT TAGCAGAACT ACACACCAGG GCCAGGGGTC AGATATCCAC

121 TGACCTTTGG ATGGTGCTAC AAGCTAGTAC CAGTTGAGCC AGATAAGGTA GAAGAGGCCA

181 ATAAAGGAGA GAACACCAGC TTGTTACACC CTGTGAGCCT GCATGGGATG GATGACCCGG

241 AGAGAGAAGT GTTAGAGTGG AGGTTTGACA GCCGCCTAGC ATTTCATCAC GTGGCCCGAG

301 AGCTGCATCC GGAGTACTTC AAGAACTGCT GATATCGAGC TTGCTACAAG GGACTTTCCG

361 CTGGGGACTT TCCAGGGAGG CGTGGCCTGG GCGGGACTGG GGAGTGGCGA GCCCTCAGAT

421 CCTGCATATA AGCAGCTGCT TTTTGCCTGT ACTGGGTCTC TCTGGTTAGA CCAGATCTGA

481 GCCTGGGAGC TCTCTGGCTA ACTAGGGAAC CCACTGCTTA AGCCTCAATA AAGCTTGCCT

541 TGAGTGCTTC AAGTAGTGTG TGCCCGTCTG TTGTGTGACT CTGGTAACTA GAGATCCCTC

601 AGACCCTTTT AGTCAGTGTG GAAAATCTCT AGCAGTGGCG CCCGAACAGG GACTTGAAAG

661 CGAAAGGGAA ACCAGAGGAG CTCTCTCGAC GCAGGACTCG GCTTGCTGAA GCGCGCACGG

721 CAAGAGGCGA GGGGCGGCGA CTGGTGAGTA CGCCAAAAAT TTTGACTAGC GGAGGCTAGA

781 AGGAGAGAGA TGGGTGCGAG AGCGTCAGTA TTAAGCGGGG GAGAATTAGA TCGCGATGGG

841 AAAAAATTCG GTTAAGGCCA GGGGGAAAGA AAAAATATAA ATTAAAACAT ATAGTATGGG

901 CAAGCAGGGA GCTAGAACGA TTCGCAGTTA ATCCTGGCCT GTTAGAAACA TCAGAAGGCT

961 GTAGACAAAT ACTGGGACAG CTACAACCAT CCCTTCAGAC AGGATCAGAA GAACTTAGAT

1021 CATTATATAA TACAGTAGCA ACCCTCTATT GTGTGCATCA AAGGATAGAG ATAAAAGACA

1081 CCAAGGAAGC TTTAGACAAG ATAGAGGAAG AGCAAAACAA AAGTAAGACC ACCGCACAGC

1141 AAGCGGCCGG CCGCTGATCT TCAGACCTGG AGGAGGAGAT ATGAGGGACA ATTGGAGAAG

1201 TGAATTATAT AAATATAAAG TAGTAAAAAT TGAACCATTA GGAGTAGCAC CCACCAAGGC

1261 AAAGAGAAGA GTGGTGCAGA GAGAAAAAAG AGCAGTGGGA ATAGGAGCTT TGTTCCTTGG

1321 GTTCTTGGGA GCAGCAGGAA GCACTATGGG CGCAGCGTCA ATGACGCTGA CGGTACAGGC

1381 CAGACAATTA TTGTCTGGTA TAGTGCAGCA GCAGAACAAT TTGCTGAGGG CTATTGAGGC

1441 GCAACAGCAT CTGTTGCAAC TCACAGTCTG GGGCATCAAG CAGCTCCAGG CAAGAATCCT

1501 GGCTGTGGAA AGATACCTAA AGGATCAACA GCTCCTGGGG ATTTGGGGTT GCTCTGGAAA

1561 ACTCATTTGC ACCACTGCTG TGCCTTGGAA TGCTAGTTGG AGTAATAAAT CTCTGGAACA