LOCUS       sequence-247139-        6488 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Puromycin resistance gene and dre recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6488)
  AUTHORS   Nam HS, Capecchi MR
  TITLE     2A plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6488)
  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            "Unpublished"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: pCAGGS-pac-T2A-dre
FEATURES             Location/Qualifiers
     source          1..6488
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        11..390
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        392..669
                     /label=chicken beta-actin promoter
     intron          670..1686
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1610..1632
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1694..1713
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1744..2340
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MAEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     2240..2260
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             2362..2415
                     /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             3478..3498
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3516..3535)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3581..3636
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3635..3654)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3997..4013)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3997..4013)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4010..4032)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4021..4037
                     /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(4045..4051,4052..4069,4070..4075))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4090..4111
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4170..4365
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4216..4351
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4371..4505
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4408..4427)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4462..4481
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4573..4590)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4744..5332)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4824..4843)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5503..6294,6295..6363))
                     /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     6126..6145
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6364..6468)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctgggtc gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
       61 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      121 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      181 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      241 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      301 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      361 atctacgtat tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact
      421 ctccccatct cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt
      481 tgtgcagcga tgggggcggg gggggggggg gggcgcgcgc caggcggggc ggggcggggc
      541 gaggggcggg gcggggcgag gcggagaggt gcggcggcag ccaatcagag cggcgcgctc
      601 cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg
      661 cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg
      721 cgccgcccgc cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc
      781 ccttctcctc cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg
      841 ctgcgtgaaa gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg
      901 ggtgcgtgcg tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc
      961 tgtgagcgct gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc
     1021 gcggccgggg gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg
     1081 cggggtgtgt gcgtgggggg gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc
     1141 cccctgcacc cccctccccg agttgctgag cacggcccgg cttcgggtgc ggggctccgt
     1201 acggggcgtg gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg
     1261 ggcggggcgg ggccgcctcg ggccggggag ggctcggggg aggggcgcgg cggcccccgg
     1321 agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc
     1381 gagagggcgc agggacttcc tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc
     1441 cgccgcaccc cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg
     1501 ggcggggagg gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg
     1561 ggctgtccgc ggggggacgg ctgccttcgg gggggacggg gcagggcggg gttcggcttc
     1621 tggcgtgtga ccggcggctc tagagcctct gctaaccatg ttcatgcctt cttctttttc
     1681 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagaattc
     1741 gccatggccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc
     1801 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg
     1861 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
     1921 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
     1981 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
     2041 ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag
     2101 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
     2161 ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc
     2221 ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc
     2281 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
     2341 ggcggctccg gcggccgctc cgagggccgc ggctccctgc tgacctgcgg cgacgtggag
     2401 gagaaccccg gccccgcccc cggctccacc atgggtgcta gcgagctgat catctctggc
     2461 tcctctggag gattcctgag gaacatcggc aaggagtacc aggaggctgc tgagaacttc
     2521 atgagattca tgaatgacca gggagcctac gcccctaaca ccctgagaga cctgaggctg
     2581 gtgttccact cctgggctag atggtgccac gctagacagc tggcctggtt ccctatctct
     2641 cctgagatgg ctagggagta cttccttcag ctgcacgatg ctgacctggc ctctaccacc
     2701 atcgacaagc actacgccat gctgaacatg ctgctgtccc actgtggcct gcctcctctg
     2761 tctgatgaca agtctgtgag cctggccatg aggagaatcc ggagagaggc tgccaccgag
     2821 aagggagaga gaaccggcca ggccatccct ctgagatggg atgacctgaa gctgctggat
     2881 gtgctgctgt ctagatctga gagactggtg gacctgagga atagggcctt cctgtttgtg
     2941 gcctacaaca ccctgatgag gatgtctgag atctctagga tcagagtggg agacctggac
     3001 cagaccggag acaccgtgac cctgcacatc tcccacacca agaccatcac caccgctgct
     3061 ggcctggaca aagtgctgtc taggaggacc accgctgtgc tgaatgactg gctggatgtg
     3121 tctggcctga gagagcaccc tgacgctgtg ctgttccctc ctatccaccg gagcaacaag
     3181 gctaggatca ccaccacccc tctgaccgcc cctgccatgg agaagatttt tagcgatgcc
     3241 tgggtgctgc tgaacaagag ggatgccacc cctaacaagg gccgctaccg gacctggacc
     3301 ggccactctg ctagagtggg agctgccatc gacatggctg agaagcaagt gtccatggtg
     3361 gagatcatgc aggagggcac ctggaaaaag cctgagacac tgatgagata cctgaggagg
     3421 ggaggagtgt ctgtgggagc caactctagg ctgatggact ccgctagcgg cgccggtcct
     3481 aagaagaaga ggaaagtgtg ataaggcgcg ccggatcttt ttccctctgc caaaaattat
     3541 ggggacatca tgaagcccct tgagcatctg acttctggct aataaaggaa atttattttc
     3601 attgcaatag tgtgttggaa ttttttgtgt ctctcactcg gaaggacata tgggagggca
     3661 aatcatttaa aacatcagaa tgagtatttg gtttagagtt tggcaacata tgcccatatg
     3721 ctggctgcca tgaacaaagg ttggctataa agaggtcatc agtatatgaa acagccccct
     3781 gctgtccatt ccttattcca tagaaaagcc ttgacttgag gttagatttt ttttatattt
     3841 tgttttgtgt tatttttttc tttaacatcc ctaaaatttt ccttacatgt tttactagcc
     3901 agatttttcc tcctctcctg actactccca gtcatagctg tccctcttct cttatggaga
     3961 tccctcgacc tgcagcccaa gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     4021 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     4081 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     4141 gtcgggaaac ctgtcgtgcc agcggatccg catctcaatt agtcagcaac catagtcccg
     4201 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     4261 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     4321 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct aacttgttta
     4381 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     4441 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     4501 ggatccgctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4561 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4621 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4681 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4741 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4801 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4861 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4921 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4981 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5041 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5101 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5161 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     5221 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5281 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     5341 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     5401 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     5461 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     5521 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5581 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5641 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5701 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5761 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5821 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5881 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5941 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     6001 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     6061 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     6121 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     6181 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     6241 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6301 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     6361 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     6421 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     6481 aaaagtgc
//
