pLVX-mCherry-C1 is an HIV-1-based, lentiviral expression vector that allows you to express your gene of interest fused to mCherry, a mutant fluorescent protein derived from the tetrameric Discosoma sp. red fluorescent protein, DsRed (1). The excitation and emission maxima of the native mCherry protein are 587 nm and 610 nm, respectively. Genes cloned into the multiple cloning site (MCS), located at the C-terminal end of the mCherry coding sequence, are expressed as C-terminal mCherry fusion proteins. Expression of the fusion protein is driven by the constitutively active human cytomegalovirus immediate early promoter (PCMV IE), located just upstream of the mCherry coding sequence. Lentiviral particles derived from the vector allow the expression of mCherry fusion proteins in virtually any cell type, including primary cells. The unmodified vector expresses mCherry, and may be used to produce marker virus to optimize infection protocols.

pLVX-mCherry-C1 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 (2), 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 (3). Finally, pLVX-mCherry-C1 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).

In addition to lentiviral elements, pLVX-mCherry-C1 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(yīng)用

To construct a fusion protein, the gene of interest must be cloned into pLVX-mCherry-C1 so that it is in-frame with the mCherry coding sequence. The inserted sequence does not require an initiation codon (ATG) or a stop codon (TAA, TAG, TGA); however, if you don't want to use the stop codons downstream of the MCS (see map), you can add a stop codon to the end of your gene of interest.

The fusion protein is constitutively expressed when pLVX-mCherry-C1 is transduced into target cells. Before the vector can be transduced, however, it must be transfected into 293T packaging cells with our Lenti-X? HT Packaging System (Cat. Nos. 632160 and 632161). 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 (5).

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 CGATAAGCTT GGGAGTTCCG CGTTACATAA CTTACGGTAA 
2221 ATGGCCCGCC TGGCTGACCG CCCAACGACC CCCGCCCATT GACGTCAATA ATGACGTATG 
2281 TTCCCATAGT AACGCCAATA GGGACTTTCC ATTGACGTCA ATGGGTGGAG TATTTACGGT 
2341 AAACTGCCCA CTTGGCAGTA CATCAAGTGT ATCATATGCC AAGTACGCCC CCTATTGACG 
2401 TCAATGACGG TAAATGGCCC GCCTGGCATT ATGCCCAGTA CATGACCTTA TGGGACTTTC 
2461 CTACTTGGCA GTACATCTAC GTATTAGTCA TCGCTATTAC CATGGTGATG CGGTTTTGGC 
2521 AGTACATCAA TGGGCGTGGA TAGCGGTTTG ACTCACGGGG ATTTCCAAGT CTCCACCCCA 
2581 TTGACGTCAA TGGGAGTTTG TTTTGGCACC AAAATCAACG GGACTTTCCA AAATGTCGTA 
2641 ACAACTCCGC CCCATTGACG CAAATGGGCG GTAGGCGTGT ACGGTGGGAG GTCTATATAA 
2701 GCAGAGCTCG TTTAGTGAAC CGTCAGATCG CCTGGAGACG CCATCCACGC TGTTTTGACC 
2761 TCCATAGAAG ACACCGACTC TACTAGAGGA TCTACCGGTC GCCACCATGG TGAGCAAGGG 
2821 CGAGGAGGAT AACATGGCCA TCATCAAGGA GTTCATGCGC TTCAAGGTGC ACATGGAGGG 
2881 CTCCGTGAAC GGCCACGAGT TCGAGATCGA GGGCGAGGGC GAGGGCCGCC CCTACGAGGG 
2941 CACCCAGACC GCCAAGCTGA AGGTGACCAA GGGTGGCCCC CTGCCCTTCG CCTGGGACAT 
3001 CCTGTCCCCT CAGTTCATGT ACGGCTCCAA GGCCTACGTG AAGCACCCCG CCGACATCCC 
3061 CGACTACTTG AAGCTGTCCT TCCCCGAGGG CTTCAAGTGG GAGCGCGTGA TGAACTTCGA 
3121 GGACGGCGGC GTGGTGACCG TGACCCAGGA CTCCTCCCTG CAGGACGGCG AGTTCATCTA 
3181 CAAGGTGAAG CTGCGCGGCA CCAACTTCCC CTCCGACGGC CCCGTAATGC AGAAGAAGAC 
3241 CATGGGCTGG GAGGCCTCCT CCGAGCGGAT GTACCCCGAG GACGGCGCCC TGAAGGGCGA 
3301 GATCAAGCAG AGGCTGAAGC TGAAGGACGG CGGCCACTAC GACGCTGAGG TCAAGACCAC 
3361 CTACAAGGCC AAGAAGCCCG TGCAGCTGCC CGGCGCCTAC AACGTCAACA TCAAGTTGGA 
3421 CATCACCTCC CACAACGAGG ACTACACCAT CGTGGAACAG TACGAACGCG CCGAGGGCCG 
3481 CCACTCCACC GGCGGCATGG ACGAGCTGTA CAAGTCCGGA CTCAGATCTC GAGCTCAAGC 
3541 TTCGAATTCT GCAGTCGACG GTACCGCGGG CCCGGGATCC ACCGGATCTA GATAACTGAT 
3601 CATAATTCTA CCGGGTAGGG GAGGCGCTTT TCCCAAGGCA GTCTGGAGCA TGCGCTTTAG 
3661 CAGCCCCGCT GGGCACTTGG CGCTACACAA GTGGCCTCTG GCCTCGCACA CATTCCACAT 
3721 CCACCGGTAG GCGCCAACCG GCTCCGTTCT TTGGTGGCCC CTTCGCGCCA CCTTCTACTC 
3781 CTCCCCTAGT CAGGAAGTTC CCCCCCGCCC CGCAGCTCGC GTCGTGCAGG ACGTGACAAA 
3841 TGGAAGTAGC ACGTCTCACT AGTCTCGTGC AGATGGACAG CACCGCTGAG CAATGGAAGC 
3901 GGGTAGGCCT TTGGGGCAGC GGCCAATAGC AGCTTTGCTC CTTCGCTTTC TGGGCTCAGA 
3961 GGCTGGGAAG GGGTGGGTCC GGGGGCGGGC TCAGGGGCGG GCTCAGGGGC GGGGCGGGCG 
4021 CCCGAAGGTC CTCCGGAGGC CCGGCATTCT GCACGCTTCA AAAGCGCACG TCTGCCGCGC 
4081 TGTTCTCCTC TTCCTCATCT CCGGGCCTTT CGACCTGCAG CCCAAGCTTA CCATGACCGA 
4141 GTACAAGCCC ACGGTGCGCC TCGCCACCCG CGACGACGTC CCCAGGGCCG TACGCACCCT 
4201 CGCCGCCGCG TTCGCCGACT ACCCCGCCAC GCGCCACACC GTCGATCCGG ACCGCCACAT 
4261 CGAGCGGGTC ACCGAGCTGC AAGAACTCTT CCTCACGCGC GTCGGGCTCG ACATCGGCAA 
4321 GGTGTGGGTC GCGGACGACG GCGCCGCGGT GGCGGTCTGG ACCACGCCGG AGAGCGTCGA 
4381 AGCGGGGGCG GTGTTCGCCG AGATCGGCCC GCGCATGGCC GAGTTGAGCG GTTCCCGGCT 
4441 GGCCGCGCAG CAACAGATGG AAGGCCTCCT GGCGCCGCAC CGGCCCAAGG AGCCCGCGTG 
4501 GTTCCTGGCC ACCGTCGGCG TCTCGCCCGA CCACCAGGGC AAGGGTCTGG GCAGCGCCGT 
4561 CGTGCTCCCC GGAGTGGAGG CGGCCGAGCG CGCCGGGGTG CCCGCCTTCC TGGAGACCTC 
4621 CGCGCCCCGC AACCTCCCCT TCTACGAGCG GCTCGGCTTC ACCGTCACCG CCGACGTCGA 
4681 GGTGCCCGAA GGACCGCGCA CCTGGTGCAT GACCCGCAAG CCCGGTGCCT GACCGCGTCT 
4741 GGAACAATCA ACCTCTGGAT TACAAAATTT GTGAAAGATT GACTGGTATT CTTAACTATG 
4801 TTGCTCCTTT TACGCTATGT GGATACGCTG CTTTAATGCC TTTGTATCAT GCTATTGCTT 
4861 CCCGTATGGC TTTCATTTTC TCCTCCTTGT ATAAATCCTG GTTGCTGTCT CTTTATGAGG 
4921 AGTTGTGGCC CGTTGTCAGG CAACGTGGCG TGGTGTGCAC TGTGTTTGCT GACGCAACCC 
4981 CCACTGGTTG GGGCATTGCC ACCACCTGTC AGCTCCTTTC CGGGACTTTC GCTTTCCCCC 
5041 TCCCTATTGC CACGGCGGAA CTCATCGCCG CCTGCCTTGC CCGCTGCTGG ACAGGGGCTC 
5101 GGCTGTTGGG CACTGACAAT TCCGTGGTGT TGTCGGGGAA GCTGACGTCC TTTCCATGGC 
5161 TGCTCGCCTG TGTTGCCACC TGGATTCTGC GCGGGACGTC CTTCTGCTAC GTCCCTTCGG 
5221 CCCTCAATCC AGCGGACCTT CCTTCCCGCG GCCTGCTGCC GGCTCTGCGG CCTCTTCCGC 
5281 GTCTTCGCCT TCGCCCTCAG ACGAGTCGGA TCTCCCTTTG GGCCGCCTCC CCGCCTGGAA 
5341 TTAATTCTGC AGTCGAGACC TAGAAAAACA TGGAGCAATC ACAAGTAGCA ATACAGCAGC 
5401 TACCAATGCT GATTGTGCCT GGCTAGAAGC ACAAGAGGAG GAGGAGGTGG GTTTTCCAGT 
5461 CACACCTCAG GTACCTTTAA GACCAATGAC TTACAAGGCA GCTGTAGATC TTAGCCACTT 
5521 TTTAAAAGAA AAGAGGGGAC TGGAAGGGCT AATTCACTCC CAACGAAGAC AAGATATCCT 
5581 TGATCTGTGG ATCTACCACA CACAAGGCTA CTTCCCTGAT TAGCAGAACT ACACACCAGG 
5641 GCCAGGGGTC AGATATCCAC TGACCTTTGG ATGGTGCTAC AAGCTAGTAC CAGTTGAGCC 
5701 AGATAAGGTA GAAGAGGCCA ATAAAGGAGA GAACACCAGC TTGTTACACC CTGTGAGCCT 
5761 GCATGGGATG GATGACCCGG AGAGAGAAGT GTTAGAGTGG AGGTTTGACA GCCGCCTAGC 
5821 ATTTCATCAC GTGGCCCGAG AGCTGCATCC GGAGTACTTC AAGAACTGCT GATATCGAGC 
5881 TTGCTACAAG GGACTTTCCG CTGGGGACTT TCCAGGGAGG CGTGGCCTGG GCGGGACTGG 
5941 GGAGTGGCGA GCCCTCAGAT CCTGCATATA AGCAGCTGCT TTTTGCCTGT ACTGGGTCTC 
6001 TCTGGTTAGA CCAGATCTGA GCCTGGGAGC TCTCTGGCTA ACTAGGGAAC CCACTGCTTA 
6061 AGCCTCAATA AAGCTTGCCT TGAGTGCTTC AAGTAGTGTG TGCCCGTCTG TTGTGTGACT 
6121 CTGGTAACTA GAGATCCCTC AGACCCTTTT AGTCAGTGTG GAAAATCTCT AGCAGTAGTA 
6181 GTTCATGTCA TCTTATTATT CAGTATTTAT AACTTGCAAA GAAATGAATA TCAGAGAGTG 
6241 AGAGGCCTTG ACATTGCTAG CGTTTTACCG TCGACCTCTA GCTAGAGCTT GGCGTAATCA 
6301 TGGTCATAGC TGTTTCCTGT GTGAAATTGT TATCCGCTCA CAATTCCACA CAACATACGA 
6361 GCCGGAAGCA TAAAGTGTAA AGCCTGGGGT GCCTAATGAG TGAGCTAACT CACATTAATT 
6421 GCGTTGCGCT CACTGCCCGC TTTCCAGTCG GGAAACCTGT CGTGCCAGCT GCATTAATGA 
6481 ATCGGCCAAC GCGCGGGGAG AGGCGGTTTG CGTATTGGGC GCTCTTCCGC TTCCTCGCTC 
6541 ACTGACTCGC TGCGCTCGGT CGTTCGGCTG CGGCGAGCGG TATCAGCTCA CTCAAAGGCG 
6601 GTAATACGGT TATCCACAGA ATCAGGGGAT AACGCAGGAA AGAACATGTG AGCAAAAGGC 
6661 CAGCAAAAGG CCAGGAACCG TAAAAAGGCC GCGTTGCTGG CGTTTTTCCA TAGGCTCCGC 
6721 CCCCCTGACG AGCATCACAA AAATCGACGC TCAAGTCAGA GGTGGCGAAA CCCGACAGGA 
6781 CTATAAAGAT ACCAGGCGTT TCCCCCTGGA AGCTCCCTCG TGCGCTCTCC TGTTCCGACC 
6841 CTGCCGCTTA CCGGATACCT GTCCGCCTTT CTCCCTTCGG GAAGCGTGGC GCTTTCTCAT 
6901 AGCTCACGCT GTAGGTATCT CAGTTCGGTG TAGGTCGTTC GCTCCAAGCT GGGCTGTGTG 
6961 CACGAACCCC CCGTTCAGCC CGACCGCTGC GCCTTATCCG GTAACTATCG TCTTGAGTCC 
7021 AACCCGGTAA GACACGACTT ATCGCCACTG GCAGCAGCCA CTGGTAACAG GATTAGCAGA 
7081 GCGAGGTATG TAGGCGGTGC TACAGAGTTC TTGAAGTGGT GGCCTAACTA CGGCTACACT 
7141 AGAAGAACAG TATTTGGTAT CTGCGCTCTG CTGAAGCCAG TTACCTTCGG AAAAAGAGTT 
7201 GGTAGCTCTT GATCCGGCAA ACAAACCACC GCTGGTAGCG GTGGTTTTTT TGTTTGCAAG 
7261 CAGCAGATTA CGCGCAGAAA AAAAGGATCT CAAGAAGATC CTTTGATCTT TTCTACGGGG 
7321 TCTGACGCTC AGTGGAACGA AAACTCACGT TAAGGGATTT TGGTCATGAG ATTATCAAAA 
7381 AGGATCTTCA CCTAGATCCT TTTAAATTAA AAATGAAGTT TTAAATCAAT CTAAAGTATA 
7441 TATGAGTAAA CTTGGTCTGA CAGTTACCAA TGCTTAATCA GTGAGGCACC TATCTCAGCG 
7501 ATCTGTCTAT TTCGTTCATC CATAGTTGCC TGACTCCCCG TCGTGTAGAT AACTACGATA 
7561 CGGGAGGGCT TACCATCTGG CCCCAGTGCT GCAATGATAC CGCGAGACCC ACGCTCACCG 
7621 GCTCCAGATT TATCAGCAAT AAACCAGCCA GCCGGAAGGG CCGAGCGCAG AAGTGGTCCT 
7681 GCAACTTTAT CCGCCTCCAT CCAGTCTATT AATTGTTGCC GGGAAGCTAG AGTAAGTAGT 
7741 TCGCCAGTTA ATAGTTTGCG CAACGTTGTT GCCATTGCTA CAGGCATCGT GGTGTCACGC 
7801 TCGTCGTTTG GTATGGCTTC ATTCAGCTCC GGTTCCCAAC GATCAAGGCG AGTTACATGA 
7861 TCCCCCATGT TGTGCAAAAA AGCGGTTAGC TCCTTCGGTC CTCCGATCGT TGTCAGAAGT 
7921 AAGTTGGCCG CAGTGTTATC ACTCATGGTT ATGGCAGCAC TGCATAATTC TCTTACTGTC 
7981 ATGCCATCCG TAAGATGCTT TTCTGTGACT GGTGAGTACT CAACCAAGTC ATTCTGAGAA 
8041 TAGTGTATGC GGCGACCGAG TTGCTCTTGC CCGGCGTCAA TACGGGATAA TACCGCGCCA 
8101 CATAGCAGAA CTTTAAAAGT GCTCATCATT GGAAAACGTT CTTCGGGGCG AAAACTCTCA 
8161 AGGATCTTAC CGCTGTTGAG ATCCAGTTCG ATGTAACCCA CTCGTGCACC CAACTGATCT 
8221 TCAGCATCTT TTACTTTCAC CAGCGTTTCT GGGTGAGCAA AAACAGGAAG GCAAAATGCC 
8281 GCAAAAAAGG GAATAAGGGC GACACGGAAA TGTTGAATAC TCATACTCTT CCTTTTTCAA 
8341 TATTATTGAA GCATTTATCA GGGTTATTGT CTCATGAGCG GATACATATT TGAATGTATT 
8401 TAGAAAAATA AACAAATAGG GGTTCCGCGC ACATTTCCCC GAAAAGTGCC ACCTGACGTC 
8461 GACGGATCGG GAGATCAACT TGTTTATTGC AGCTTATAAT GGTTACAAAT AAAGCAATAG 
8521 CATCACAAAT TTCACAAATA AAGCATTTTT TTCACTGCAT TCTAGTTGTG GTTTGTCCAA 
8581 ACTCATCAAT GTATCTTATC ATGTCTGGAT CAACTGGATA ACTCAAGCTA ACCAAAATCA 
8641 TCCCAAACTT CCCACCCCAT ACCCTATTAC CACTGCCAAT TACCTGTGGT TTCATTTACT 
8701 CTAAACCTGT GATTCCTCTG AATTATTTTC ATTTTAAAGA AATTGTATTT GTTAAATATG 
8761 TACTACAAAC TTAGTAGT