pLVX-Tight-Puro is a tetracycline (Tet)-inducible, lentiviral expression vector designed to express a gene of interest under the control of PTight, a modified Tet-responsive promoter. PTight consists of a modified minimal CMV promoter, and seven direct repeats of a 36 bp regulatory sequence that contains the 19 bp tet operator sequence (tetO; 1). This vector is designed to be used with our Lenti-XTM Tet-On? Advanced and Tet-Off? Advanced Inducible Expression Systems (Cat. Nos. 632162 and 632163). These systems provide the inducible gene expression strategy of Gossen & Bujard, with major improvements described by Urlinger, et al. (2-6), in a lentiviral format.

pLVX-Tight-Puro 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 enhances nuclear export of viral and transgene RNA (7), 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 (8). Finally, pLVX-Tight-Puro 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 (9)..

In addition to lentiviral elements, pLVX-Tight-Puro 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.

載體應用

pLVX-Tight-Puro is a lentiviral expression vector that allows tightly regulated, doxycycline-controlled expression of a gene of interest. In order to function, the system requires the presence of a tetracycline-controlled transcriptional activator (rtTA Advanced or tTA Advanced) supplied by either pLVX-Tet-On Advanced or pLVX-Tet-off Advanced lentiviruses. Before pLVX-Tight-Puro can be transduced into cells, 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 infectious, replication-incompetent, VSV-G pseudotyped lentiviral particles that can infect a wide range of cell types, including non-dividing and primary cells (10).

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 CGATGAGGCC CTTTCGTCTT CACTCGAGTT TACTCCCTAT 
2221 CAGTGATAGA GAACGTATGT CGAGTTTACT CCCTATCAGT GATAGAGAAC GATGTCGAGT 
2281 TTACTCCCTA TCAGTGATAG AGAACGTATG TCGAGTTTAC TCCCTATCAG TGATAGAGAA 
2341 CGTATGTCGA GTTTACTCCC TATCAGTGAT AGAGAACGTA TGTCGAGTTT ATCCCTATCA 
2401 GTGATAGAGA ACGTATGTCG AGTTTACTCC CTATCAGTGA TAGAGAACGT ATGTCGAGGT 
2461 AGGCGTGTAC GGTGGGAGGC CTATATAAGC AGAGCTCGTT TAGTGAACCG TCAGATCGCC 
2521 TGGAGAAGGA TCCGCGGCCG CGCCGGCTCT AGATCGCGAA CGCGTGAATT CTACCGGGTA 
2581 GGGGAGGCGC TTTTCCCAAG GCAGTCTGGA GCATGCGCTT TAGCAGCCCC GCTGGGCACT 
2641 TGGCGCTACA CAAGTGGCCT CTGGCCTCGC ACACATTCCA CATCCACCGG TAGGCGCCAA 
2701 CCGGCTCCGT TCTTTGGTGG CCCCTTCGCG CCACCTTCTA CTCCTCCCCT AGTCAGGAAG 
2761 TTCCCCCCCG CCCCGCAGCT CGCGTCGTGC AGGACGTGAC AAATGGAAGT AGCACGTCTC 
2821 ACTAGTCTCG TGCAGATGGA CAGCACCGCT GAGCAATGGA AGCGGGTAGG CCTTTGGGGC 
2881 AGCGGCCAAT AGCAGCTTTG CTCCTTCGCT TTCTGGGCTC AGAGGCTGGG AAGGGGTGGG 
2941 TCCGGGGGCG GGCTCAGGGG CGGGCTCAGG GGCGGGGCGG GCGCCCGAAG GTCCTCCGGA 
3001 GGCCCGGCAT TCTGCACGCT TCAAAAGCGC ACGTCTGCCG CGCTGTTCTC CTCTTCCTCA 
3061 TCTCCGGGCC TTTCGACCTG CAGCCCAAGC TTACCATGAC CGAGTACAAG CCCACGGTGC 
3121 GCCTCGCCAC CCGCGACGAC GTCCCCAGGG CCGTACGCAC CCTCGCCGCC GCGTTCGCCG 
3181 ACTACCCCGC CACGCGCCAC ACCGTCGATC CGGACCGCCA CATCGAGCGG GTCACCGAGC 
3241 TGCAAGAACT CTTCCTCACG CGCGTCGGGC TCGACATCGG CAAGGTGTGG GTCGCGGACG 
3301 ACGGCGCCGC GGTGGCGGTC TGGACCACGC CGGAGAGCGT CGAAGCGGGG GCGGTGTTCG 
3361 CCGAGATCGG CCCGCGCATG GCCGAGTTGA GCGGTTCCCG GCTGGCCGCG CAGCAACAGA 
3421 TGGAAGGCCT CCTGGCGCCG CACCGGCCCA AGGAGCCCGC GTGGTTCCTG GCCACCGTCG 
3481 GCGTCTCGCC CGACCACCAG GGCAAGGGTC TGGGCAGCGC CGTCGTGCTC CCCGGAGTGG 
3541 AGGCGGCCGA GCGCGCCGGG GTGCCCGCCT TCCTGGAGAC CTCCGCGCCC CGCAACCTCC 
3601 CCTTCTACGA GCGGCTCGGC TTCACCGTCA CCGCCGACGT CGAGGTGCCC GAAGGACCGC 
3661 GCACCTGGTG CATGACCCGC AAGCCCGGTG CCTGACGGGC GCGTCTGGAA CAATCAACCT 
3721 CTGGATTACA AAATTTGTGA AAGATTGACT GGTATTCTTA ACTATGTTGC TCCTTTTACG 
3781 CTATGTGGAT ACGCTGCTTT AATGCCTTTG TATCATGCTA TTGCTTCCCG TATGGCTTTC 
3841 ATTTTCTCCT CCTTGTATAA ATCCTGGTTG CTGTCTCTTT ATGAGGAGTT GTGGCCCGTT 
3901 GTCAGGCAAC GTGGCGTGGT GTGCACTGTG TTTGCTGACG CAACCCCCAC TGGTTGGGGC 
3961 ATTGCCACCA CCTGTCAGCT CCTTTCCGGG ACTTTCGCTT TCCCCCTCCC TATTGCCACG 
4021 GCGGAACTCA TCGCCGCCTG CCTTGCCCGC TGCTGGACAG GGGCTCGGCT GTTGGGCACT 
4081 GACAATTCCG TGGTGTTGTC GGGGAAGCTG ACGTCCTTTC CATGGCTGCT CGCCTGTGTT 
4141 GCCACCTGGA TTCTGCGCGG GACGTCCTTC TGCTACGTCC CTTCGGCCCT CAATCCAGCG 
4201 GACCTTCCTT CCCGCGGCCT GCTGCCGGCT CTGCGGCCTC TTCCGCGTCT TCGCCTTCGC 
4261 CCTCAGACGA GTCGGATCTC CCTTTGGGCC GCCTCCCCGC CTGGAATTAA TTCTGCAGTC 
4321 GAGACCTAGA AAAACATGGA GCAATCACAA GTAGCAATAC AGCAGCTACC AATGCTGATT 
4381 GTGCCTGGCT AGAAGCACAA GAGGAGGAGG AGGTGGGTTT TCCAGTCACA CCTCAGGTAC 
4441 CTTTAAGACC AATGACTTAC AAGGCAGCTG TAGATCTTAG CCACTTTTTA AAAGAAAAGA 
4501 GGGGACTGGA AGGGCTAATT CACTCCCAAC GAAGACAAGA TATCCTTGAT CTGTGGATCT 
4561 ACCACACACA AGGCTACTTC CCTGATTAGC AGAACTACAC ACCAGGGCCA GGGGTCAGAT 
4621 ATCCACTGAC CTTTGGATGG TGCTACAAGC TAGTACCAGT TGAGCCAGAT AAGGTAGAAG 
4681 AGGCCAATAA AGGAGAGAAC ACCAGCTTGT TACACCCTGT GAGCCTGCAT GGGATGGATG 
4741 ACCCGGAGAG AGAAGTGTTA GAGTGGAGGT TTGACAGCCG CCTAGCATTT CATCACGTGG 
4801 CCCGAGAGCT GCATCCGGAG TACTTCAAGA ACTGCTGATA TCGAGCTTGC TACAAGGGAC 
4861 TTTCCGCTGG GGACTTTCCA GGGAGGCGTG GCCTGGGCGG GACTGGGGAG TGGCGAGCCC 
4921 TCAGATCCTG CATATAAGCA GCTGCTTTTT GCCTGTACTG GGTCTCTCTG GTTAGACCAG 
4981 ATCTGAGCCT GGGAGCTCTC TGGCTAACTA GGGAACCCAC TGCTTAAGCC TCAATAAAGC 
5041 TTGCCTTGAG TGCTTCAAGT AGTGTGTGCC CGTCTGTTGT GTGACTCTGG TAACTAGAGA 
5101 TCCCTCAGAC CCTTTTAGTC AGTGTGGAAA ATCTCTAGCA GTAGTAGTTC ATGTCATCTT 
5161 ATTATTCAGT ATTTATAACT TGCAAAGAAA TGAATATCAG AGAGTGAGAG GCCTTGACAT 
5221 TGCTAGCGTT TACCGTCGAC CTCTAGCTAG AGCTTGGCGT AATCATGGTC ATAGCTGTTT 
5281 CCTGTGTGAA ATTGTTATCC GCTCACAATT CCACACAACA TACGAGCCGG AAGCATAAAG 
5341 TGTAAAGCCT GGGGTGCCTA ATGAGTGAGC TAACTCACAT TAATTGCGTT GCGCTCACTG 
5401 CCCGCTTTCC AGTCGGGAAA CCTGTCGTGC CAGCTGCATT AATGAATCGG CCAACGCGCG 
5461 GGGAGAGGCG GTTTGCGTAT TGGGCGCTCT TCCGCTTCCT CGCTCACTGA CTCGCTGCGC 
5521 TCGGTCGTTC GGCTGCGGCG AGCGGTATCA GCTCACTCAA AGGCGGTAAT ACGGTTATCC 
5581 ACAGAATCAG GGGATAACGC AGGAAAGAAC ATGTGAGCAA AAGGCCAGCA AAAGGCCAGG 
5641 AACCGTAAAA AGGCCGCGTT GCTGGCGTTT TTCCATAGGC TCCGCCCCCC TGACGAGCAT 
5701 CACAAAAATC GACGCTCAAG TCAGAGGTGG CGAAACCCGA CAGGACTATA AAGATACCAG 
5761 GCGTTTCCCC CTGGAAGCTC CCTCGTGCGC TCTCCTGTTC CGACCCTGCC GCTTACCGGA 
5821 TACCTGTCCG CCTTTCTCCC TTCGGGAAGC GTGGCGCTTT CTCATAGCTC ACGCTGTAGG 
5881 TATCTCAGTT CGGTGTAGGT CGTTCGCTCC AAGCTGGGCT GTGTGCACGA ACCCCCCGTT 
5941 CAGCCCGACC GCTGCGCCTT ATCCGGTAAC TATCGTCTTG AGTCCAACCC GGTAAGACAC 
6001 GACTTATCGC CACTGGCAGC AGCCACTGGT AACAGGATTA GCAGAGCGAG GTATGTAGGC 
6061 GGTGCTACAG AGTTCTTGAA GTGGTGGCCT AACTACGGCT ACACTAGAAG AACAGTATTT 
6121 GGTATCTGCG CTCTGCTGAA GCCAGTTACC TTCGGAAAAA GAGTTGGTAG CTCTTGATCC 
6181 GGCAAACAAA CCACCGCTGG TAGCGGTGGT TTTTTTGTTT GCAAGCAGCA GATTACGCGC 
6241 AGAAAAAAAG GATCTCAAGA AGATCCTTTG ATCTTTTCTA CGGGGTCTGA CGCTCAGTGG 
6301 AACGAAAACT CACGTTAAGG GATTTTGGTC ATGAGATTAT CAAAAAGGAT CTTCACCTAG 
6361 ATCCTTTTAA ATTAAAAATG AAGTTTTAAA TCAATCTAAA GTATATATGA GTAAACTTGG 
6421 TCTGACAGTT ACCAATGCTT AATCAGTGAG GCACCTATCT CAGCGATCTG TCTATTTCGT 
6481 TCATCCATAG TTGCCTGACT CCCCGTCGTG TAGATAACTA CGATACGGGA GGGCTTACCA 
6541 TCTGGCCCCA GTGCTGCAAT GATACCGCGA GACCCACGCT CACCGGCTCC AGATTTATCA 
6601 GCAATAAACC AGCCAGCCGG AAGGGCCGAG CGCAGAAGTG GTCCTGCAAC TTTATCCGCC 
6661 TCCATCCAGT CTATTAATTG TTGCCGGGAA GCTAGAGTAA GTAGTTCGCC AGTTAATAGT 
6721 TTGCGCAACG TTGTTGCCAT TGCTACAGGC ATCGTGGTGT CACGCTCGTC GTTTGGTATG 
6781 GCTTCATTCA GCTCCGGTTC CCAACGATCA AGGCGAGTTA CATGATCCCC CATGTTGTGC 
6841 AAAAAAGCGG TTAGCTCCTT CGGTCCTCCG ATCGTTGTCA GAAGTAAGTT GGCCGCAGTG 
6901 TTATCACTCA TGGTTATGGC AGCACTGCAT AATTCTCTTA CTGTCATGCC ATCCGTAAGA 
6961 TGCTTTTCTG TGACTGGTGA GTACTCAACC AAGTCATTCT GAGAATAGTG TATGCGGCGA 
7021 CCGAGTTGCT CTTGCCCGGC GTCAATACGG GATAATACCG CGCCACATAG CAGAACTTTA 
7081 AAAGTGCTCA TCATTGGAAA ACGTTCTTCG GGGCGAAAAC TCTCAAGGAT CTTACCGCTG 
7141 TTGAGATCCA GTTCGATGTA ACCCACTCGT GCACCCAACT GATCTTCAGC ATCTTTTACT 
7201 TTCACCAGCG TTTCTGGGTG AGCAAAAACA GGAAGGCAAA ATGCCGCAAA AAAGGGAATA 
7261 AGGGCGACAC GGAAATGTTG AATACTCATA CTCTTCCTTT TTCAATATTA TTGAAGCATT 
7321 TATCAGGGTT ATTGTCTCAT GAGCGGATAC ATATTTGAAT GTATTTAGAA AAATAAACAA 
7381 ATAGGGGTTC CGCGCACATT TCCCCGAAAA GTGCCACCTG ACGTCGACGG ATCGGGAGAT 
7441 CAACTTGTTT ATTGCAGCTT ATAATGGTTA CAAATAAAGC AATAGCATCA CAAATTTCAC 
7501 AAATAAAGCA TTTTTTTCAC TGCATTCTAG TTGTGGTTTG TCCAAACTCA TCAATGTATC 
7561 TTATCATGTC TGGATCAACT GGATAACTCA AGCTAACCAA AATCATCCCA AACTTCCCAC 
7621 CCCATACCCT ATTACCACTG CCAATTACCT GTGGTTTCAT TTACTCTAAA CCTGTGATTC 
7681 CTCTGAATTA TTTTCATTTT AAAGAAATTG TATTTGTTAA ATATGTACTA CAAACTTAGT 
7741 AGTTTTTAAA GAAATTGTAT TTGTTAAATA TGTACTACAA ACTTAGTAGT