LOCUS       sequence-177025-        5240 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  PV4 for 2-input, 4-output decoder circuit.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5240)
  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 5240)
  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: pBW770_pCAG-PV4-loxP-mRuby2-BGHpA
FEATURES             Location/Qualifiers
     source          1..5240
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     346..368
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     430..449
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(548..581)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      591..600
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             597..1307
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     polyA_signal    1311..1535
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1781..1800)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    1846..1901
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(1900..1919)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2262..2278)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2262..2278)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2275..2297)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2286..2302
                     /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(2310..2316,2317..2334,2335..2340))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2355..2376
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2461..2595
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2615..2749
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2652..2671)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2706..2725
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2817..2834)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2988..3576)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3068..3087)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3747..4538,4539..4607))
                     /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     4370..4389
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4608..4712)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4743..5122
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc ccccggagcg
       61 ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa tcgtgcgaga
      121 gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg aggcgccgcc
      181 gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag gaaatgggcg
      241 gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag cctcggggct
      301 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
      361 gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac
      421 agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattgatta
      481 attcgagcga aggcgcgccc tgacctcctg ccagcaatag taagacaaca cgcaaagtcg
      541 cacgcgtata acttcgtata gcatacatta tacgaagtta taccggtgcc gccaccatgg
      601 tgtctaaggg cgaagagctg atcaaggaaa atatgcgtat gaaggtggtc atggaaggtt
      661 cggtcaacgg ccaccaattc aaatgcacag gtgaaggaga aggcaatccg tacatgggaa
      721 ctcaaaccat gaggatcaaa gtcatcgagg gaggacccct gccatttgcc tttgacattc
      781 ttgccacgtc gttcatgtat ggcagccgta cttttatcaa gtacccgaaa ggcattcctg
      841 atttctttaa acagtccttt cctgagggtt ttacttggga aagagttacg agatacgaag
      901 atggtggagt cgtcaccgtc atgcaggaca ccagccttga ggatggctgt ctcgtttacc
      961 acgtccaagt cagaggggta aactttccct ccaatggtcc cgtgatgcag aagaagacca
     1021 agggttggga gcctaataca gagatgatgt atccagcaga tggtggtctg aggggataca
     1081 ctcatatggc actgaaagtt gatggtggtg gccatctgtc ttgctctttc gtaacaactt
     1141 acaggtcaaa aaagaccgtc gggaacatca agatgcccgg tatccatgcc gttgatcacc
     1201 gcctggaaag gttagaggaa agtgacaatg aaatgttcgt agtacaacgc gaacacgcag
     1261 ttgccaagtt cgccgggctt ggtggtggga tggacgagct gtacaagtaa ctgtgccttc
     1321 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     1381 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     1441 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     1501 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
     1561 gggctctagg gggtatcccc acgcgccctg tagcggaatt cggtaccgag ccaactccct
     1621 ttacaacctc actcaagtcc gttagaggcg gccgcccagg atacatagat taccacaact
     1681 ccgagccctt ccaccgctag cgcactcctc aggtgcaggc tgcctatcag aaggtggtgg
     1741 ctggtgtggc caatgccctg gctcacaaat accactgaga tctttttccc tctgccaaaa
     1801 attatgggga catcatgaag ccccttgagc atctgacttc tggctaataa aggaaattta
     1861 ttttcattgc aatagtgtgt tggaattttt tgtgtctctc actcggaagg acatatggga
     1921 gggcaaatca tttaaaacat cagaatgagt atttggttta gagtttggca acatatgccc
     1981 atatgctggc tgccatgaac aaaggttggc tataaagagg tcatcagtat atgaaacagc
     2041 cccctgctgt ccattcctta ttccatagaa aagccttgac ttgaggttag atttttttta
     2101 tattttgttt tgtgttattt ttttctttaa catccctaaa attttcctta catgttttac
     2161 tagccagatt tttcctcctc tcctgactac tcccagtcat agctgtccct cttctcttat
     2221 ggagatccct cgacctgcag cccaagcttg gcgtaatcat ggtcatagct gtttcctgtg
     2281 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     2341 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     2401 ttccagtcgg gaaacctgtc gtgccagcgg atccgcatct caattagtca gcaaccatag
     2461 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     2521 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     2581 gcttttttgg aggcctaggc ttttgcaaaa agctaacttg tttattgcag cttataatgg
     2641 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     2701 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatcc gctgcattaa
     2761 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     2821 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     2881 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     2941 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     3001 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     3061 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     3121 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     3181 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     3241 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     3301 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     3361 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     3421 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     3481 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     3541 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     3601 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     3661 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     3721 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     3781 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     3841 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     3901 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     3961 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     4021 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     4081 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     4141 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     4201 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     4261 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     4321 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     4381 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     4441 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     4501 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     4561 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     4621 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     4681 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctggg
     4741 tcgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
     4801 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     4861 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     4921 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     4981 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     5041 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     5101 attagtcatc gctattacca tggtcgaggt gagccccacg ttctgcttca ctctccccat
     5161 ctcccccccc tccccacccc caattttgta tttatttatt ttttaattat tttgtgcagc
     5221 gatgggggcg gggggggggg
//
