LOCUS       sequence-177151-        5621 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  PV3 for 2-input, 4-output decoder circuit.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5621)
  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 5621)
  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: pBW769_pCAG-PV3-FRT-loxP-iRFP720-BGHpA
FEATURES             Location/Qualifiers
     source          1..5621
                     /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=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      746..755
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             752..1702
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 720 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP720
                     /note="derived from Rhodopseudomonas palustris RpBphP2"
                     /translation="MAEGSVARQPDLLTCDDEPIHIPGAIQPHGLLLALAADMTIVAGS
                     DNLPELTGLAIGALIGRSAADVFDSETHNRLTIALAEPGAAVGAPITVGFTMRKDAGFI
                     GSWHRHDQLIFLELEPPQRDVAEPQAFFRRTNSAIRRLQAAETLESACAAAAQEVRKIT
                     GFDRVMIYRFASDFSGSVIAEDRCAEVESKLGLHYPASFIPAQARRLYTINPVRIIPDI
                     NYRPVPVTPDLNPVTGRPIDLSFAILRSVSPNHLEFMRNIGMHGTMSISILRGERLWGL
                     IVCHHRTPYYVDLDGRQACELVAQVLAWQIGVMEE"
     polyA_signal    1703..1927
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2173..2192)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2238..2293
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2292..2311)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2654..2670)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2654..2670)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2667..2689)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2678..2694
                     /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(2702..2708,2709..2726,2727..2732))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2747..2768
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2853..2987
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3007..3141
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3044..3063)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3098..3117
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3209..3226)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3380..3968)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3460..3479)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4139..4930,4931..4999))
                     /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     4762..4781
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5000..5104)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        5135..5514
                     /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 cactgaaggt cctcaatcgc actggaaaca tcaaggtcgg
      661 cacgcgtgaa gttcctattc tctagaaagt ataggaactt cgataacttc gtatagcata
      721 cattatacga agttataccg gtgccgccac catggcggaa ggatccgtcg ccaggcagcc
      781 tgacctcttg acctgcgacg atgagccgat ccatatcccc ggtgccatcc aaccgcatgg
      841 actgctgctc gccctcgccg ccgacatgac gatcgttgcc ggcagcgaca accttcccga
      901 actcaccgga ctggcgatcg gcgccctgat cggccgctct gcggccgatg tcttcgactc
      961 ggagacgcac aaccgtctga cgatcgcctt ggccgagccc ggggcggccg tcggagcacc
     1021 gatcactgtc ggcttcacga tgcgaaagga cgcaggcttc atcggctcct ggcatcgcca
     1081 tgatcagctc atcttcctcg agctcgagcc tccccagcgg gacgtcgccg agccgcaggc
     1141 gttcttccgc cgcaccaaca gcgccatccg ccgcctgcag gccgccgaaa ccttggaaag
     1201 cgcctgcgcc gccgcggcgc aagaggtgcg gaagattacc ggcttcgatc gggtgatgat
     1261 ctatcgcttc gcctccgact tcagcgggtc cgtgatcgca gaggatcggt gcgccgaggt
     1321 cgagtcaaaa ctaggcctgc actatcctgc ctcattcatc ccggcgcagg cccgtcggct
     1381 ctataccatc aacccggtac ggatcattcc cgatatcaat tatcggccgg tgccggtcac
     1441 cccagacctc aatccggtca ccgggcggcc gattgatctt agcttcgcca tcctgcgcag
     1501 cgtctcgccc aaccatctgg agttcatgcg caacataggc atgcacggca cgatgtcgat
     1561 ctcgattttg cgcggcgagc gactgtgggg attgatcgtt tgccatcacc gaacgccgta
     1621 ctacgtcgat ctcgatggcc gccaagcctg cgagctagtc gcccaggttc tggcctggca
     1681 gatcggcgtg atggaagagt gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     1741 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     1801 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     1861 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     1921 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     1981 tgtagcggaa ttcggtaccc tgacctcctg ccagcaatag taagacaaca cgcaaagtcg
     2041 cggccgccca ggatacatag attaccacaa ctccgagccc ttccaccgct agcgcactcc
     2101 tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc tggctcacaa
     2161 ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga agccccttga
     2221 gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt gttggaattt
     2281 tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac atcagaatga
     2341 gtatttggtt tagagtttgg caacatatgc ccatatgctg gctgccatga acaaaggttg
     2401 gctataaaga ggtcatcagt atatgaaaca gccccctgct gtccattcct tattccatag
     2461 aaaagccttg acttgaggtt agattttttt tatattttgt tttgtgttat ttttttcttt
     2521 aacatcccta aaattttcct tacatgtttt actagccaga tttttcctcc tctcctgact
     2581 actcccagtc atagctgtcc ctcttctctt atggagatcc ctcgacctgc agcccaagct
     2641 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     2701 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     2761 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     2821 ggatccgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccagttccgc
     2881 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     2941 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     3001 aaagctaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     3061 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     3121 gtatcttatc atgtctggat ccgctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     3181 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     3241 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     3301 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     3361 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     3421 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     3481 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     3541 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     3601 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     3661 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     3721 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     3781 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     3841 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3901 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     3961 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     4021 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     4081 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     4141 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     4201 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     4261 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     4321 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     4381 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     4441 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     4501 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     4561 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     4621 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     4681 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     4741 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     4801 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     4861 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     4921 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     4981 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     5041 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     5101 cgcgcacatt tccccgaaaa gtgccacctg ggtcgacatt gattattgac tagttattaa
     5161 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
     5221 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
     5281 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
     5341 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
     5401 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
     5461 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtcgag
     5521 gtgagcccca cgttctgctt cactctcccc atctcccccc cctccccacc cccaattttg
     5581 tatttattta ttttttaatt attttgtgca gcgatggggg c
//
