LOCUS       sequence-177043-        5596 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  PV1 for 3-input, 2-output Full Adder.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5596)
  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 5596)
  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: pBW758_pCAG-PV1-VloxP-FRT-lox2272-NeoR-BGHpA
FEATURES             Location/Qualifiers
     source          1..5596
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     347..369
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     431..450
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    583..616
                     /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(617..650)
                     /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             699..1493
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(753..772)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1363..1382
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1667..1891
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2137..2156)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2202..2257
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2256..2275)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2618..2634)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2618..2634)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2631..2653)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2642..2658
                     /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(2666..2672,2673..2690,2691..2696))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2711..2732
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2817..2951
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2971..3105
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3008..3027)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3062..3081
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3173..3190)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3344..3932)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3424..3443)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4103..4894,4895..4963))
                     /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     4726..4745
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4964..5068)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        5099..5478
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ggggcggggc cgcctcgggc cggggagggc tcgggggagg ggcgcggcgg cccccggagc
       61 gccggcggct gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag
      121 agggcgcagg gacttccttt gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc
      181 cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc
      241 ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc tccctctcca gcctcggggc
      301 tgtccgcggg gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg
      361 cgtgtgaccg gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta
      421 cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattgatt
      481 aattcgagcg aaggcgcgcc cattactcgc atccattctc aggctgtctc gtctcgtctc
      541 gcacgcgttc aatttctgag aactgtcatt ctcggaaatt gagaagttcc tattctctag
      601 aaagtatagg aacttcataa cttcgtatag gatactttat acgaagttat accggtcgga
      661 tttgatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
      721 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
      781 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
      841 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
      901 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
      961 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     1021 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     1081 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     1141 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     1201 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     1261 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     1321 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     1381 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     1441 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     1501 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     1561 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     1621 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccctgt gccttctagt
     1681 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     1741 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     1801 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     1861 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc
     1921 tctagggggt atccccacgc gccctgtagc ggaattcggt accgctggga gttcgtagac
     1981 ggaaacaaac gcagaatcca agcgcggccg cccaggatac atagattacc acaactccga
     2041 gcccttccac cgctagcgca ctcctcaggt gcaggctgcc tatcagaagg tggtggctgg
     2101 tgtggccaat gccctggctc acaaatacca ctgagatctt tttccctctg ccaaaaatta
     2161 tggggacatc atgaagcccc ttgagcatct gacttctggc taataaagga aatttatttt
     2221 cattgcaata gtgtgttgga attttttgtg tctctcactc ggaaggacat atgggagggc
     2281 aaatcattta aaacatcaga atgagtattt ggtttagagt ttggcaacat atgcccatat
     2341 gctggctgcc atgaacaaag gttggctata aagaggtcat cagtatatga aacagccccc
     2401 tgctgtccat tccttattcc atagaaaagc cttgacttga ggttagattt tttttatatt
     2461 ttgttttgtg ttattttttt ctttaacatc cctaaaattt tccttacatg ttttactagc
     2521 cagatttttc ctcctctcct gactactccc agtcatagct gtccctcttc tcttatggag
     2581 atccctcgac ctgcagccca agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     2641 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     2701 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     2761 agtcgggaaa cctgtcgtgc cagcggatcc gcatctcaat tagtcagcaa ccatagtccc
     2821 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     2881 ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt
     2941 ttttggaggc ctaggctttt gcaaaaagct aacttgttta ttgcagctta taatggttac
     3001 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     3061 tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatccgctg cattaatgaa
     3121 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     3181 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     3241 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     3301 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     3361 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     3421 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     3481 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     3541 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     3601 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     3661 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     3721 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     3781 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     3841 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     3901 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     3961 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     4021 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     4081 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     4141 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     4201 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     4261 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     4321 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     4381 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     4441 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     4501 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     4561 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     4621 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     4681 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     4741 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     4801 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     4861 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     4921 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     4981 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     5041 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgggtcga
     5101 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
     5161 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
     5221 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
     5281 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
     5341 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
     5401 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
     5461 gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct ccccatctcc
     5521 cccccctccc cacccccaat tttgtattta tttatttttt aattattttg tgcagcgatg
     5581 ggggcggggg gggggg
//
