LOCUS       sequence-322454-        7611 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mCherry and puromycin resistance gene under CMV promoter. 
            3rd gen lentiviral vector..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7611)
  TITLE     lenti vector
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7611)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pCDH-CMV-mCherry-T2A-Puro
FEATURES             Location/Qualifiers
     source          1..7611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     180..202
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     194..211
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     195..211
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        226..452
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             453..633
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    680..805
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1298..1531
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1716..1760
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2027..2143
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2233..2436
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2387..2407
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2433..2457
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2521..3228
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2666..2684)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2934..2953
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3244..3297
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3298..3897
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3298..3317)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3794..3814
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    3910..4498
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3963..3983)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4381..4392)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4572..4805
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4877..4998
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4914..4933)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4968..4987
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5038..5173
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5206..5222)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5206..5222)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5219..5241)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5230..5246
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5254..5260,5261..5278,5279..5284))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5299..5320
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5437..5454)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5608..6196)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6367..7158,7159..7227))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6990..7009
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7228..7332)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7400..7418
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7456..7478)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7578..7597
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
       61 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
      121 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
      181 ccagtcacga cgttgtaaaa cgacggccag tgccaagctg acgcgtgtag tcttatgcaa
      241 tactcttgta gtcttgcaac atggtaacga tgagttagca acatgcctta caaggagaga
      301 aaaagcaccg tgcatgccga ttggtggaag taaggtggta cgatcgtgcc ttattaggaa
      361 ggcaacagac gggtctgaca tggattggac gaaccactga attgccgcat tgcagagata
      421 ttgtatttaa gtgcctagct cgatacataa acgggtctct ctggttagac cagatctgag
      481 cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt
      541 gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca
      601 gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc
      661 gaaagggaaa ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc
      721 aagaggcgag gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa
      781 ggagagagat gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga
      841 aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc
      901 aagcagggag ctagaacgat tcgcagttaa tcctggcctg ttagaaacat cagaaggctg
      961 tagacaaata ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc
     1021 attatataat acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac
     1081 caaggaagct ttagacaaga tagaggaaga gcaaaacaaa agtaagacca ccgcacagca
     1141 agcggccact gatcttcaga cctggaggag gagatatgag ggacaattgg agaagtgaat
     1201 tatataaata taaagtagta aaaattgaac cattaggagt agcacccacc aaggcaaaga
     1261 gaagagtggt gcagagagaa aaaagagcag tgggaatagg agctttgttc cttgggttct
     1321 tgggagcagc aggaagcact atgggcgcag cgtcaatgac gctgacggta caggccagac
     1381 aattattgtc tggtatagtg cagcagcaga acaatttgct gagggctatt gaggcgcaac
     1441 agcatctgtt gcaactcaca gtctggggca tcaagcagct ccaggcaaga atcctggctg
     1501 tggaaagata cctaaaggat caacagctcc tggggatttg gggttgctct ggaaaactca
     1561 tttgcaccac tgctgtgcct tggaatgcta gttggagtaa taaatctctg gaacagattt
     1621 ggaatcacac gacctggatg gagtgggaca gagaaattaa caattacaca agcttaatac
     1681 actccttaat tgaagaatcg caaaaccagc aagaaaagaa tgaacaagaa ttattggaat
     1741 tagataaatg ggcaagtttg tggaattggt ttaacataac aaattggctg tggtatataa
     1801 aattattcat aatgatagta ggaggcttgg taggtttaag aatagttttt gctgtacttt
     1861 ctatagtgaa tagagttagg cagggatatt caccattatc gtttcagacc cacctcccaa
     1921 ccccgagggg acccgacagg cccgaaggaa tagaagaaga aggtggagag agagacagag
     1981 acagatccat tcgattagtg aacggatctc gacggtatcg gttaactttt aaaagaaaag
     2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2101 aaactaaaga attacaaaaa caaattacaa aattcaaaat tttatcgata ctagtattat
     2161 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
     2221 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
     2281 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     2341 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
     2401 aggcgtgtac ggtgggaggt ttatataagc agagctcgtt tagtgaaccg tcagatcgcc
     2461 tggagacgcc atccacgctg ttttgacctc catagaagat tctagagcta gggatccacc
     2521 atggtgagca agggcgagga ggataacatg gccatcatca aggagttcat gcgcttcaag
     2581 gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc
     2641 cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc
     2701 ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta cgtgaagcac
     2761 cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa gtgggagcgc
     2821 gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac
     2881 ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta
     2941 atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc
     3001 gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca ctacgacgct
     3061 gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc
     3121 aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgaa
     3181 cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagaa gcttggcagt
     3241 ggagagggca gaggaagtct gctaacatgc ggtgacgtcg aggagaatcc tggcccaatg
     3301 accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc
     3361 accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga tccggaccgc
     3421 cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc
     3481 ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac gccggagagc
     3541 gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt gagcggttcc
     3601 cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc caaggagccc
     3661 gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg tctgggcagc
     3721 gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag
     3781 acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt caccgccgac
     3841 gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg tgcctgagaa
     3901 ttcgtcgaca atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     3961 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     4021 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     4081 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     4141 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     4201 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     4261 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct
     4321 tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     4381 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     4441 ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcct
     4501 ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga
     4561 aaagggggga ctggaagggc taattcactc ccaacgaaga taagatctgc tttttgcttg
     4621 tactgggtct ctctggttag accagatctg agcctgggag ctctctggct aactagggaa
     4681 cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct
     4741 gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc
     4801 tagcagtagt agttcatgtc atcttattat tcagtattta taacttgcaa agaaatgaat
     4861 atcagagagt gagaggaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4921 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4981 actcatcaat gtatcttatc atgtctggct ctagctatcc cgcccctaac tccgcccatc
     5041 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     5101 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
     5161 ttttttggag gcctagactt ttgcagagac caaattcgta atcatgtcat agctgtttcc
     5221 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     5281 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     5341 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5401 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     5461 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5521 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5581 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5641 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5701 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5761 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5821 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5881 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5941 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     6001 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     6061 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     6121 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     6181 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     6241 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     6301 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     6361 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     6421 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6481 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6541 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6601 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6661 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6721 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6781 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6841 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6901 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6961 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     7021 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     7081 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     7141 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     7201 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     7261 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     7321 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat
     7381 catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg cgcgtttcgg
     7441 tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta
     7501 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg
     7561 gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc a
//
