LOCUS       sequence-177158-        5334 bp    DNA     linear   UNA 14-APR-2025
DEFINITION  Expresses VCre under pCAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5334)
  AUTHORS   Weinberg BH, Pham NT, Caraballo LD, Lozanoski T, Engel A, Bhatia S, 
            Wong WW
  TITLE     Large-scale design of robust genetic circuits with multiple inputs 
            and outputs for mammalian cells.
  JOURNAL   Nat Biotechnol. 2017 Mar 27. doi: 10.1038/nbt.3805.
  PUBMED    28346402
REFERENCE   2  (bases 1 to 5334)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2017 Mar 27. doi: 10.1038/nbt.3805."
COMMENT     Sequence Label: pCAG-VCre
FEATURES             Location/Qualifiers
     source          1..5334
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     466..488
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     550..569
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             629..649
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      1677..1686
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1875..1894)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    1940..1995
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(1994..2013)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2356..2372)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2356..2372)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2369..2391)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2380..2396
                     /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(2404..2410,2411..2428,2429..2434))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2449..2470
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2555..2689
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2709..2843
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2746..2765)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2800..2819
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2911..2928)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3082..3670)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3162..3181)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3841..4632,4633..4701))
                     /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     4464..4483
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4702..4806)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4837..5216
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg ctccgtacgg
       61 ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg gtgccgggcg
      121 gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc ccccggagcg
      181 ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa tcgtgcgaga
      241 gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg aggcgccgcc
      301 gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag gaaatgggcg
      361 gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag cctcggggct
      421 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
      481 gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac
      541 agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattgatta
      601 attcgagcga acgcgtgccg ccaccatgcc caagaaaaag cggaaagtga tcgagaacca
      661 gctgagcctg ctgggcgact tttctggcgt gcggcccgac gatgtgaaaa ccgccattca
      721 ggccgcccag aaaaagggca tcaacgtggc cgagaacgag cagttcaagg ccgccttcga
      781 gcatctgctg aacgagttca agaagcggga agagagatac agccccaaca ccctgcggcg
      841 gctggaaagc gcctggacct gcttcgtgga ttggtgcctg gccaaccaca gacacagcct
      901 gcctgccacc cccgataccg tggaagcctt cttcatcgag cgggccgagg aactgcaccg
      961 gaacaccctg agcgtgtaca gatgggccat cagccgggtg cacagagtgg ccggatgccc
     1021 tgatccctgc ctggacatct acgtggaaga tcggctgaag gccattgccc ggaagaaagt
     1081 gcgggaaggc gaggccgtga agcaggccag ccctttcaac gagcagcatc tgctgaagct
     1141 gaccagcctg tggtacagaa gcgacaagct gctgctgcgg cggaacctgg ctctgctggc
     1201 tgtggcctac gagagcatgc tgagagccag cgagctggcc aacatccggg tgtccgatat
     1261 ggaactggcc ggcgacggaa ccgccatcct gaccatccct atcaccaaga ccaaccactc
     1321 cggcgagccc gatacctgca tcctgtccca ggatgtggtg tccctgctga tggactacac
     1381 cgaggccggc aagctggata tgagcagcga cggcttcctg ttcgtgggcg tgtccaagca
     1441 caacacctgt atcaagccca agaaggacaa gcagaccggc gaggtgctgc acaagcccat
     1501 caccaccaag acagtggaag gcgtgttcta cagcgcctgg gagacactgg acctgggcag
     1561 acagggcgtg aagcctttca cagcccacag cgccagagtg ggagccgctc aggacctgct
     1621 gaagaagggc tacaataccc tgcagatcca gcagtccggc cggtggtcta gcggagccat
     1681 ggtggccaga tacggcagag ccatcctggc tagggatggc gctatggccc acagcagagt
     1741 gaaaaccaga tccgccccca tgcagtgggg caaggacgag aaggactgag cggccgcact
     1801 cctcaggtgc aggctgccta tcagaaggtg gtggctggtg tggccaatgc cctggctcac
     1861 aaataccact gagatctttt tccctctgcc aaaaattatg gggacatcat gaagcccctt
     1921 gagcatctga cttctggcta ataaaggaaa tttattttca ttgcaatagt gtgttggaat
     1981 tttttgtgtc tctcactcgg aaggacatat gggagggcaa atcatttaaa acatcagaat
     2041 gagtatttgg tttagagttt ggcaacatat gcccatatgc tggctgccat gaacaaaggt
     2101 tggctataaa gaggtcatca gtatatgaaa cagccccctg ctgtccattc cttattccat
     2161 agaaaagcct tgacttgagg ttagattttt tttatatttt gttttgtgtt atttttttct
     2221 ttaacatccc taaaattttc cttacatgtt ttactagcca gatttttcct cctctcctga
     2281 ctactcccag tcatagctgt ccctcttctc ttatggagat ccctcgacct gcagcccaag
     2341 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     2401 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     2461 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     2521 gcggatccgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccagttcc
     2581 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     2641 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     2701 aaaaagctaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2761 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     2821 atgtatctta tcatgtctgg atccgctgca ttaatgaatc ggccaacgcg cggggagagg
     2881 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     2941 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     3001 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     3061 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     3121 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     3181 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     3241 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     3301 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     3361 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     3421 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     3481 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     3541 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     3601 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     3661 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     3721 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3781 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3841 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     3901 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     3961 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     4021 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     4081 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     4141 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     4201 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     4261 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     4321 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     4381 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     4441 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     4501 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     4561 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     4621 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     4681 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     4741 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     4801 tccgcgcaca tttccccgaa aagtgccacc tgggtcgaca ttgattattg actagttatt
     4861 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
     4921 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
     4981 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
     5041 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
     5101 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
     5161 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg
     5221 aggtgagccc cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt
     5281 tgtatttatt tattttttaa ttattttgtg cagcgatggg ggcggggggg gggg
//
