LOCUS       sequence-177149-        5421 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  PV2 for 2-input, 4-output decoder circuit and 3-input, 2-output Full
            Adder.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5421)
  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 5421)
  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: "Nat 
            Biotechnol. 2017 Mar 27. doi: 10.1038/nbt.3805."
COMMENT     Sequence Label: pBW759_pCAG-PV2-lox2272-GFP-BGHpA
FEATURES             Location/Qualifiers
     source          1..5421
                     /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"
     protein_bind    complement(668..701)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     regulatory      711..720
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(717..719,720..722,723..1436)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(717..719,720..722,723..1433)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(762..783)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1023..1042)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1370..1391
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1437..1661
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1907..1926)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    1972..2027
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2026..2045)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2388..2404)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2388..2404)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2401..2423)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2412..2428
                     /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(2436..2442,2443..2460,2461..2466))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2481..2502
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2587..2721
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2741..2875
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2778..2797)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2832..2851
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2943..2960)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3114..3702)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3194..3213)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3873..4664,4665..4733))
                     /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     4496..4515
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4734..4838)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4869..5248
                     /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 aggcgcgccg ctgggagttc gtagacggaa acaaacgcag aatccaagcg
      661 cacgcgtata acttcgtata ggatacttta tacgaagtta taccggtgcc gccaccatgg
      721 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
      781 acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca
      841 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
      901 tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc
      961 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca
     1021 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
     1081 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
     1141 tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca
     1201 tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc
     1261 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     1321 tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
     1381 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtaactgt
     1441 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     1501 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     1561 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     1621 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     1681 cagctggggc tctagggggt atccccacgc gccctgtagc ggaattcggt accgcactga
     1741 aggtcctcaa tcgcactgga aacatcaagg tcggcggccg cccaggatac atagattacc
     1801 acaactccga gcccttccac cgctagcgca ctcctcaggt gcaggctgcc tatcagaagg
     1861 tggtggctgg tgtggccaat gccctggctc acaaatacca ctgagatctt tttccctctg
     1921 ccaaaaatta tggggacatc atgaagcccc ttgagcatct gacttctggc taataaagga
     1981 aatttatttt cattgcaata gtgtgttgga attttttgtg tctctcactc ggaaggacat
     2041 atgggagggc aaatcattta aaacatcaga atgagtattt ggtttagagt ttggcaacat
     2101 atgcccatat gctggctgcc atgaacaaag gttggctata aagaggtcat cagtatatga
     2161 aacagccccc tgctgtccat tccttattcc atagaaaagc cttgacttga ggttagattt
     2221 tttttatatt ttgttttgtg ttattttttt ctttaacatc cctaaaattt tccttacatg
     2281 ttttactagc cagatttttc ctcctctcct gactactccc agtcatagct gtccctcttc
     2341 tcttatggag atccctcgac ctgcagccca agcttggcgt aatcatggtc atagctgttt
     2401 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     2461 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     2521 cccgctttcc agtcgggaaa cctgtcgtgc cagcggatcc gcatctcaat tagtcagcaa
     2581 ccatagtccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     2641 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     2701 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct aacttgttta ttgcagctta
     2761 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     2821 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatccgctg
     2881 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     2941 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3001 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3061 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3121 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3181 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3241 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3301 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3361 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3421 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3481 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     3541 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     3601 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     3661 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     3721 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     3781 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     3841 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     3901 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     3961 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4021 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4081 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4141 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4201 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4261 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     4321 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     4381 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     4441 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     4501 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     4561 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     4621 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     4681 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     4741 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     4801 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     4861 cctgggtcga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
     4921 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     4981 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     5041 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     5101 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     5161 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     5221 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
     5281 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
     5341 tgcagcgatg ggggcggggg gggggggggc gcgcgccagg cggggcgggg cggggcgagg
     5401 ggcggggcgg ggcgaggcgg a
//
