pLVX-shRNA2 is an HIV-1-based, lentiviral expression vector designed to express a small hairpin RNA (shRNA) for RNA interference (RNAi) studies. Expression of your shRNA is driven by the RNA Pol III-dependent, human U6 promoter (PU6), located just upstream of the MCS. pLVX-shRNA2 can be used as a plasmid expression vector and transfected into cells, or it can be packaged into viral particles and transduced into cells. Lentiviral particles derived from the vector allow the expression of shRNAs in virtually any cell type, including primary cells.

In addition to expressing shRNAs, pLVX-shRNA2 also expresses the fluorescent protein ZsGreen1, a human codon-optimized variant of the reef coral Zoanthus sp. green fluorescent protein, ZsGreen (1). Expression of ZsGreen1 is driven by the constitutively active human cytomegalovirus immediate early promoter (PCMV IE), allowing it to be used as an indicator of transfection or transduction efficiency, as well as a marker for cell sorting.

pLVX-shRNA2 contains all of the viral processing elements necessary for the production of replication-incompetent lentivirus, as well as elements to improve viral titer and overall vector function. The woodchuck hepatitis virus posttranscriptional regulatory element (WPRE) promotes RNA processing events and enhances nuclear export of viral RNA (2), leading to increased viral titers from packaging 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 (3). Finally, pLVX-shRNA2 also contains a central polypurine tract/central termination sequence element (cPPT/CTS). During target cell infection, this element creates a central DNA flap that increases nuclear import of the viral genome, resulting in improved vector integration and more efficient transduction (4). The vector also contains a pUC origin of replication and an E. coli ampicillin resistance gene (Ampr) for propagation and selection in bacteria.

載體應(yīng)用

Before pLVX-shRNA2 can be transduced into target cells, the vector must be cotransfected into 293T cells with our Lenti-X? HTX Packaging System (Cat. Nos. 631247 and 631249) and packaged into viral particles.This packaging system allows the safe production of 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 (5).

ZsGreen1 is the brightest commercially available green fluorescent protein. The presence of this protein allows transductants to be sorted by flow cytometry with standard FITC filter sets (ZsGreen1 has an excitation maximum of 493 nm and an emission maximum of 505 nm).

0001 TGGAAGGGCT AATTCACTCC CAAAGAAGAC AAGATATCCT TGATCTGTGG ATCTACCACA 
0061 CACAAGGCTA CTTCCCTGAT TAGCAGAACT ACACACCAGG GCCAGGGGTC AGATATCCAC 
0121 TGACCTTTGG ATGGTGCTAC AAGCTAGTAC CAGTTGAGCC AGATAAGGTA GAAGAGGCCA 
0181 ATAAAGGAGA GAACACCAGC TTGTTACACC CTGTGAGCCT GCATGGGATG GATGACCCGG 
0241 AGAGAGAAGT GTTAGAGTGG AGGTTTGACA GCCGCCTAGC ATTTCATCAC GTGGCCCGAG 
0301 AGCTGCATCC GGAGTACTTC AAGAACTGCT GATATCGAGC TTGCTACAAG GGACTTTCCG 
0361 CTGGGGACTT TCCAGGGAGG CGTGGCCTGG GCGGGACTGG GGAGTGGCGA GCCCTCAGAT 
0421 CCTGCATATA AGCAGCTGCT TTTTGCCTGT ACTGGGTCTC TCTGGTTAGA CCAGATCTGA 
0481 GCCTGGGAGC TCTCTGGCTA ACTAGGGAAC CCACTGCTTA AGCCTCAATA AAGCTTGCCT 
0541 TGAGTGCTTC AAGTAGTGTG TGCCCGTCTG TTGTGTGACT CTGGTAACTA GAGATCCCTC 
0601 AGACCCTTTT AGTCAGTGTG GAAAATCTCT AGCAGTGGCG CCCGAACAGG GACTTGAAAG 
0661 CGAAAGGGAA ACCAGAGGAG CTCTCTCGAC GCAGGACTCG GCTTGCTGAA GCGCGCACGG 
0721 CAAGAGGCGA GGGGCGGCGA CTGGTGAGTA CGCCAAAAAT TTTGACTAGC GGAGGCTAGA 
0781 AGGAGAGAGA TGGGTGCGAG AGCGTCAGTA TTAAGCGGGG GAGAATTAGA TCGCGATGGG 
0841 AAAAAATTCG GTTAAGGCCA GGGGGAAAGA AAAAATATAA ATTAAAACAT ATAGTATGGG 
0901 CAAGCAGGGA GCTAGAACGA TTCGCAGTTA ATCCTGGCCT GTTAGAAACA TCAGAAGGCT 
0961 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 
1621 GATTTGGAAT CACACGACCT GGATGGAGTG GGACAGAGAA ATTAACAATT ACACAAGCTT 
1681 AATACACTCC TTAATTGAAG AATCGCAAAA CCAGCAAGAA AAGAATGAAC AAGAATTATT 
1741 GGAATTAGAT AAATGGGCAA GTTTGTGGAA TTGGTTTAAC ATAACAAATT GGCTGTGGTA 
1801 TATAAAATTA TTCATAATGA TAGTAGGAGG CTTGGTAGGT TTAAGAATAG TTTTTGCTGT 
1861 ACTTTCTATA GTGAATAGAG TTAGGCAGGG ATATTCACCA TTATCGTTTC AGACCCACCT 
1921 CCCAACCCCG AGGGGACCCG ACAGGCCCGA AGGAATAGAA GAAGAAGGTG GAGAGAGAGA 
1981 CAGAGACAGA TCCATTCGAT TAGTGAACGG ATCTCGACGG TATCGCCTTT AAAAGAAAAG 
2041 GGGGGATTGG GGGGTACAGT GCAGGGGAAA GAATAGTAGA CATAATAGCA ACAGACATAC 
2101 AAACTAAAGA ATTACAAAAA CAAATTACAA AAATTCAAAA TTTTCGGGTT TATTACAGGG 
2161 ACAGCAGAGA TCCAGTTTAT CGATCTGGGC AGGAAGAGGG CCTATTTCCC ATGATTCCTT 
2221 CATATTTGCA TATACGATAC AAGGCTGTTA GAGAGATAAT TAGAATTAAT TTGACTGTAA 
2281 ACACAAAGAT ATTAGTACAA AATACGTGAC GTAGAAAGTA ATAATTTCTT GGGTAGTTTG 
2341 CAGTTTTAAA ATTATGTTTT AAAATGGACT ATCATATGCT TACCGTAACT TGAAAGTATT 
2401 TCGATTTCTT GGCTTTATAT ATCTTGTGGA AAGGACGAGG ATCCGGACAA GCTTCGAATT 
2461 CTAGTTATTA ATAGTAATCA ATTACGGGGT CATTAGTTCA TAGCCCATAT ATGGAGTTCC 
2521 GCGTTACATA ACTTACGGTA AATGGCCCGC CTGGCTGACC GCCCAACGAC CCCCGCCCAT 
2581 TGACGTCAAT AATGACGTAT GTTCCCATAG TAACGCCAAT AGGGACTTTC CATTGACGTC 
2641 AATGGGTGGA GTATTTACGG TAAACTGCCC ACTTGGCAGT ACATCAAGTG TATCATATGC 
2701 CAAGTACGCC CCCTATTGAC GTCAATGACG GTAAATGGCC CGCCTGGCAT TATGCCCAGT 
2761 ACATGACCTT ATGGGACTTT CCTACTTGGC AGTACATCTA CGTATTAGTC ATCGCTATTA 
2821 CCATGGTGAT GCGGTTTTGG CAGTACATCA ATGGGCGTGG ATAGCGGTTT GACTCACGGG 
2881 GATTTCCAAG TCTCCACCCC ATTGACGTCA ATGGGAGTTT GTTTTGGCAC CAAAATCAAC 
2941 GGGACTTTCC AAAATGTCGT AACAACTCCG CCCCATTGAC GCAAATGGGC GGTAGGCGTG 
3001 TACGGTGGGA GGTCTATATA AGCAGAGCTG GTTTAGTGAA CCGTCAGATC CGCTAGCGCT 
3061 ACCGGTCGCC ACCATGGCCC AGTCCAAGCA CGGCCTGACC AAGGAGATGA CCATGAAGTA 
3121 CCGCATGGAG GGCTGCGTGG ACGGCCACAA GTTCGTGATC ACCGGCGAGG GCATCGGCTA 
3181 CCCCTTCAAG GGCAAGCAGG CCATCAACCT GTGCGTGGTG GAGGGCGGCC CCTTGCCCTT 
3241 CGCCGAGGAC ATCTTGTCCG CCGCCTTCAT GTACGGCAAC CGCGTGTTCA CCGAGTACCC 
3301 CCAGGACATC GTCGACTACT TCAAGAACTC CTGCCCCGCC GGCTACACCT GGGACCGCTC 
3361 CTTCCTGTTC GAGGACGGCG CCGTGTGCAT CTGCAACGCC GACATCACCG TGAGCGTGGA 
3421 GGAGAACTGC ATGTACCACG AGTCCAAGTT CTACGGCGTG AACTTCCCCG CCGACGGCCC 
3481 CGTGATGAAG AAGATGACCG ACAACTGGGA GCCCTCCTGC GAGAAGATCA TCCCCGTGCC 
3541 CAAGCAGGGC ATCTTGAAGG GCGACGTGAG CATGTACCTG CTGCTGAAGG ACGGTGGCCG 
3601 CTTGCGCTGC CAGTTCGACA CCGTGTACAA GGCCAAGTCC GTGCCCCGCA AGATGCCCGA 
3661 CTGGCACTTC ATCCAGCACA AGCTGACCCG CGAGGACCGC AGCGACGCCA AGAACCAGAA 
3721 GTGGCACCTG ACCGAGCACG CCATCGCCTC CGGCTCCGCC TTGCCCTAAC TCGAGTCTAG 
3781 AGGGCCCGAC GCGTCTGGAA CAATCAACCT CTGGATTACA AAATTTGTGA AAGATTGACT 
3841 GGTATTCTTA ACTATGTTGC TCCTTTTACG CTATGTGGAT ACGCTGCTTT AATGCCTTTG 
3901 TATCATGCTA TTGCTTCCCG TATGGCTTTC ATTTTCTCCT CCTTGTATAA ATCCTGGTTG 
3961 CTGTCTCTTT ATGAGGAGTT GTGGCCCGTT GTCAGGCAAC GTGGCGTGGT GTGCACTGTG 
4021 TTTGCTGACG CAACCCCCAC TGGTTGGGGC ATTGCCACCA CCTGTCAGCT CCTTTCCGGG 
4081 ACTTTCGCTT TCCCCCTCCC TATTGCCACG GCGGAACTCA TCGCCGCCTG CCTTGCCCGC 
4141 TGCTGGACAG GGGCTCGGCT GTTGGGCACT GACAATTCCG TGGTGTTGTC GGGGAAGCTG 
4201 ACGTCCTTTC CATGGCTGCT CGCCTGTGTT GCCACCTGGA TTCTGCGCGG GACGTCCTTC 
4261 TGCTACGTCC CTTCGGCCCT CAATCCAGCG GACCTTCCTT CCCGCGGCCT GCTGCCGGCT 
4321 CTGCGGCCTC TTCCGCGTCT TCGCCTTCGC CCTCAGACGA GTCGGATCTC CCTTTGGGCC 
4381 GCCTCCCCGC CTGGAATTAA TTCTGCAGTC GAGACCTAGA AAAACATGGA GCAATCACAA 
4441 GTAGCAATAC AGCAGCTACC AATGCTGATT GTGCCTGGCT AGAAGCACAA GAGGAGGAGG 
4501 AGGTGGGTTT TCCAGTCACA CCTCAGGTAC CTTTAAGACC AATGACTTAC AAGGCAGCTG 
4561 TAGATCTTAG CCACTTTTTA AAAGAAAAGA GGGGACTGGA AGGGCTAATT CACTCCCAAC 
4621 GAAGACAAGA TATCCTTGAT CTGTGGATCT ACCACACACA AGGCTACTTC CCTGATTAGC 
4681 AGAACTACAC ACCAGGGCCA GGGGTCAGAT ATCCACTGAC CTTTGGATGG TGCTACAAGC 
4741 TAGTACCAGT TGAGCCAGAT AAGGTAGAAG AGGCCAATAA AGGAGAGAAC ACCAGCTTGT 
4801 TACACCCTGT GAGCCTGCAT GGGATGGATG ACCCGGAGAG AGAAGTGTTA GAGTGGAGGT 
4861 TTGACAGCCG CCTAGCATTT CATCACGTGG CCCGAGAGCT GCATCCGGAG TACTTCAAGA 
4921 ACTGCTGATA TCGAGCTTGC TACAAGGGAC TTTCCGCTGG GGACTTTCCA GGGAGGCGTG 
4981 GCCTGGGCGG GACTGGGGAG TGGCGAGCCC TCAGATCCTG CATATAAGCA GCTGCTTTTT 
5041 GCCTGTACTG GGTCTCTCTG GTTAGACCAG ATCTGAGCCT GGGAGCTCTC TGGCTAACTA 
5101 GGGAACCCAC TGCTTAAGCC TCAATAAAGC TTGCCTTGAG TGCTTCAAGT AGTGTGTGCC 
5161 CGTCTGTTGT GTGACTCTGG TAACTAGAGA TCCCTCAGAC CCTTTTAGTC AGTGTGGAAA 
5221 ATCTCTAGCA GTAGTAGTTC ATGTCATCTT ATTATTCAGT ATTTATAACT TGCAAAGAAA 
5281 TGAATATCAG AGAGTGAGAG GCCTTGACAT TGCTAGCGTT TACCGTCGAC CTCTAGCTAG 
5341 AGCTTGGCGT AATCATGGTC ATAGCTGTTT CCTGTGTGAA ATTGTTATCC GCTCACAATT 
5401 CCACACAACA TACGAGCCGG AAGCATAAAG TGTAAAGCCT GGGGTGCCTA ATGAGTGAGC 
5461 TAACTCACAT TAATTGCGTT GCGCTCACTG CCCGCTTTCC AGTCGGGAAA CCTGTCGTGC 
5521 CAGCTGCATT AATGAATCGG CCAACGCGCG GGGAGAGGCG GTTTGCGTAT TGGGCGCTCT 
5581 TCCGCTTCCT CGCTCACTGA CTCGCTGCGC TCGGTCGTTC GGCTGCGGCG AGCGGTATCA 
5641 GCTCACTCAA AGGCGGTAAT ACGGTTATCC ACAGAATCAG GGGATAACGC AGGAAAGAAC 
5701 ATGTGAGCAA AAGGCCAGCA AAAGGCCAGG AACCGTAAAA AGGCCGCGTT GCTGGCGTTT 
5761 TTCCATAGGC TCCGCCCCCC TGACGAGCAT CACAAAAATC GACGCTCAAG TCAGAGGTGG 
5821 CGAAACCCGA CAGGACTATA AAGATACCAG GCGTTTCCCC CTGGAAGCTC CCTCGTGCGC 
5881 TCTCCTGTTC CGACCCTGCC GCTTACCGGA TACCTGTCCG CCTTTCTCCC TTCGGGAAGC 
5941 GTGGCGCTTT CTCATAGCTC ACGCTGTAGG TATCTCAGTT CGGTGTAGGT CGTTCGCTCC 
6001 AAGCTGGGCT GTGTGCACGA ACCCCCCGTT CAGCCCGACC GCTGCGCCTT ATCCGGTAAC 
6061 TATCGTCTTG AGTCCAACCC GGTAAGACAC GACTTATCGC CACTGGCAGC AGCCACTGGT 
6121 AACAGGATTA GCAGAGCGAG GTATGTAGGC GGTGCTACAG AGTTCTTGAA GTGGTGGCCT 
6181 AACTACGGCT ACACTAGAAG AACAGTATTT GGTATCTGCG CTCTGCTGAA GCCAGTTACC 
6241 TTCGGAAAAA GAGTTGGTAG CTCTTGATCC GGCAAACAAA CCACCGCTGG TAGCGGTGGT 
6301 TTTTTTGTTT GCAAGCAGCA GATTACGCGC AGAAAAAAAG GATCTCAAGA AGATCCTTTG 
6361 ATCTTTTCTA CGGGGTCTGA CGCTCAGTGG AACGAAAACT CACGTTAAGG GATTTTGGTC 
6421 ATGAGATTAT CAAAAAGGAT CTTCACCTAG ATCCTTTTAA ATTAAAAATG AAGTTTTAAA 
6481 TCAATCTAAA GTATATATGA GTAAACTTGG TCTGACAGTT ACCAATGCTT AATCAGTGAG 
6541 GCACCTATCT CAGCGATCTG TCTATTTCGT TCATCCATAG TTGCCTGACT CCCCGTCGTG 
6601 TAGATAACTA CGATACGGGA GGGCTTACCA TCTGGCCCCA GTGCTGCAAT GATACCGCGA 
6661 GACCCACGCT CACCGGCTCC AGATTTATCA GCAATAAACC AGCCAGCCGG AAGGGCCGAG 
6721 CGCAGAAGTG GTCCTGCAAC TTTATCCGCC TCCATCCAGT CTATTAATTG TTGCCGGGAA 
6781 GCTAGAGTAA GTAGTTCGCC AGTTAATAGT TTGCGCAACG TTGTTGCCAT TGCTACAGGC 
6841 ATCGTGGTGT CACGCTCGTC GTTTGGTATG GCTTCATTCA GCTCCGGTTC CCAACGATCA 
6901 AGGCGAGTTA CATGATCCCC CATGTTGTGC AAAAAAGCGG TTAGCTCCTT CGGTCCTCCG 
6961 ATCGTTGTCA GAAGTAAGTT GGCCGCAGTG TTATCACTCA TGGTTATGGC AGCACTGCAT 
7021 AATTCTCTTA CTGTCATGCC ATCCGTAAGA TGCTTTTCTG TGACTGGTGA GTACTCAACC 
7081 AAGTCATTCT GAGAATAGTG TATGCGGCGA CCGAGTTGCT CTTGCCCGGC GTCAATACGG 
7141 GATAATACCG CGCCACATAG CAGAACTTTA AAAGTGCTCA TCATTGGAAA ACGTTCTTCG 
7201 GGGCGAAAAC TCTCAAGGAT CTTACCGCTG TTGAGATCCA GTTCGATGTA ACCCACTCGT 
7261 GCACCCAACT GATCTTCAGC ATCTTTTACT TTCACCAGCG TTTCTGGGTG AGCAAAAACA 
7321 GGAAGGCAAA ATGCCGCAAA AAAGGGAATA AGGGCGACAC GGAAATGTTG AATACTCATA 
7381 CTCTTCCTTT TTCAATATTA TTGAAGCATT TATCAGGGTT ATTGTCTCAT GAGCGGATAC 
7441 ATATTTGAAT GTATTTAGAA AAATAAACAA ATAGGGGTTC CGCGCACATT TCCCCGAAAA 
7501 GTGCCACCTG ACGTCGACGG ATCGGGAGAT CAACTTGTTT ATTGCAGCTT ATAATGGTTA 
7561 CAAATAAAGC AATAGCATCA CAAATTTCAC AAATAAAGCA TTTTTTTCAC TGCATTCTAG 
7621 TTGTGGTTTG TCCAAACTCA TCAATGTATC TTATCATGTC TGGATCAACT GGATAACTCA 
7681 AGCTAACCAA AATCATCCCA AACTTCCCAC CCCATACCCT ATTACCACTG CCAATTACCT 
7741 GTGGTTTCAT TTACTCTAAA CCTGTGATTC CTCTGAATTA TTTTCATTTT AAAGAAATTG 
7801 TATTTGTTAA ATATGTACTA CAAACTTAGT AGTTTTTAAA GAAATTGTAT TTGTTAAATA 
7861 TGTACTACAA ACTTAGTAGT