LOCUS       sequence-177155-        5390 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  PV1 for 2-input, 4-output decoder circuit.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5390)
  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 5390)
  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: pBW768_pCAG-PV1-FRT-lox2272-BFP-BGHpA
FEATURES             Location/Qualifiers
     source          1..5390
                     /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    668..701
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    complement(703..736)
                     /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)."
     CDS             752..1453
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     polyA_signal    1454..1678
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1924..1943)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    1989..2044
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2043..2062)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2405..2421)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2405..2421)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2418..2440)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2429..2445
                     /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(2453..2459,2460..2477,2478..2483))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2498..2519
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2604..2738
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2758..2892
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2795..2814)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2849..2868
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2960..2977)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3131..3719)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3211..3230)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3890..4681,4682..4750))
                     /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     4513..4532
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4751..4855)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4886..5265
                     /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 aggcgcgccc attactcgca tccattctca ggctgtctcg tctcgtctcg
      661 cacgcgtgaa gttcctattc tctagaaagt ataggaactt cgataacttc gtataggata
      721 ctttatacga agttataccg gtgccgccac catgagcgag ctgattaagg agaacatgca
      781 catgaagctg tacatggagg gcaccgtgga caaccatcac ttcaagtgca catccgaggg
      841 cgaaggcaag ccctacgagg gcacccagac catgagaatc aaggtggtcg agggcggccc
      901 tctccccttc gccttcgaca tcctggctac tagcttcctc tacggcagca agaccttcat
      961 caaccacacc cagggcatcc ccgacttctt caagcagtcc ttccctgagg gcttcacatg
     1021 ggagagagtc accacatacg aagacggggg cgtgctgacc gctacccagg acaccagcct
     1081 ccaggacggc tgcctcatct acaacgtcaa gatcagaggg gtgaacttca catccaacgg
     1141 ccctgtgatg cagaagaaaa cactcggctg ggaggccttc accgagacgc tgtaccccgc
     1201 tgacggcggc ctggaaggca gaaacgacat ggccctgaag ctcgtgggcg ggagccatct
     1261 gatcgcaaac atcaagacca catatagatc caagaaaccc gctaagaacc tcaagatgcc
     1321 tggcgtctac tatgtggact acagactgga aagaatcaag gaggccaaca acgagaccta
     1381 cgtcgagcag cacgaggtgg cagtggccag atactgcgac ctccctagca aactggggca
     1441 caagcttaat taactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     1501 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     1561 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     1621 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     1681 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcgga
     1741 attcggtacc gctgggagtt cgtagacgga aacaaacgca gaatccaagc gcggccgccc
     1801 aggatacata gattaccaca actccgagcc cttccaccgc tagcgcactc ctcaggtgca
     1861 ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc ctggctcaca aataccactg
     1921 agatcttttt ccctctgcca aaaattatgg ggacatcatg aagccccttg agcatctgac
     1981 ttctggctaa taaaggaaat ttattttcat tgcaatagtg tgttggaatt ttttgtgtct
     2041 ctcactcgga aggacatatg ggagggcaaa tcatttaaaa catcagaatg agtatttggt
     2101 ttagagtttg gcaacatatg cccatatgct ggctgccatg aacaaaggtt ggctataaag
     2161 aggtcatcag tatatgaaac agccccctgc tgtccattcc ttattccata gaaaagcctt
     2221 gacttgaggt tagatttttt ttatattttg ttttgtgtta tttttttctt taacatccct
     2281 aaaattttcc ttacatgttt tactagccag atttttcctc ctctcctgac tactcccagt
     2341 catagctgtc cctcttctct tatggagatc cctcgacctg cagcccaagc ttggcgtaat
     2401 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     2461 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     2521 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgca
     2581 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccagttccg cccattctcc
     2641 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     2701 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctaac
     2761 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     2821 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     2881 catgtctgga tccgctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     2941 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3001 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3061 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3121 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3181 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3241 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3301 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3361 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3421 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     3481 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     3541 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     3601 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     3661 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     3721 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     3781 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     3841 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     3901 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     3961 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4021 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4081 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4141 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4201 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4261 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4321 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4381 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     4441 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     4501 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     4561 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     4621 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     4681 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     4741 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     4801 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     4861 ttccccgaaa agtgccacct gggtcgacat tgattattga ctagttatta atagtaatca
     4921 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     4981 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     5041 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     5101 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     5161 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     5221 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtcga ggtgagcccc
     5281 acgttctgct tcactctccc catctccccc ccctccccac ccccaatttt gtatttattt
     5341 attttttaat tattttgtgc agcgatgggg gcgggggggg gggggcgcgc
//
