LOCUS       sequence-177150-        5401 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  PV3 for 3-input, 2-output Full Adder.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5401)
  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 5401)
  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: pBW806_pCAG-PV3-FRT-loxP-GFP-BGHpA
FEATURES             Location/Qualifiers
     source          1..5401
                     /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             join(752..754,755..757,758..1471)
                     /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(752..754,755..757,758..1468)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(797..818)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1058..1077)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1405..1426
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1472..1696
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1942..1961)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2007..2062
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2061..2080)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2423..2439)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2423..2439)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2436..2458)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2447..2463
                     /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(2471..2477,2478..2495,2496..2501))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2516..2537
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2622..2756
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2776..2910
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2813..2832)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2867..2886
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2978..2995)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3149..3737)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3229..3248)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3908..4699,4700..4768))
                     /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     4531..4550
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4769..4873)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4904..5283
                     /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 catggtgagc aagggcgagg agctgttcac
      781 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
      841 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
      901 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
      961 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     1021 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     1081 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     1141 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     1201 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     1261 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     1321 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa
     1381 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     1441 cactctcggc atggacgagc tgtacaagta actgtgcctt ctagttgcca gccatctgtt
     1501 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     1561 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     1621 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     1681 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     1741 cacgcgccct gtagcggaat tcggtaccct gacctcctgc cagcaatagt aagacaacac
     1801 gcaaagtcgc ggccgcccag gatacataga ttaccacaac tccgagccct tccaccgcta
     1861 gcgcactcct caggtgcagg ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct
     1921 ggctcacaaa taccactgag atctttttcc ctctgccaaa aattatgggg acatcatgaa
     1981 gccccttgag catctgactt ctggctaata aaggaaattt attttcattg caatagtgtg
     2041 ttggaatttt ttgtgtctct cactcggaag gacatatggg agggcaaatc atttaaaaca
     2101 tcagaatgag tatttggttt agagtttggc aacatatgcc catatgctgg ctgccatgaa
     2161 caaaggttgg ctataaagag gtcatcagta tatgaaacag ccccctgctg tccattcctt
     2221 attccataga aaagccttga cttgaggtta gatttttttt atattttgtt ttgtgttatt
     2281 tttttcttta acatccctaa aattttcctt acatgtttta ctagccagat ttttcctcct
     2341 ctcctgacta ctcccagtca tagctgtccc tcttctctta tggagatccc tcgacctgca
     2401 gcccaagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     2461 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     2521 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     2581 cgtgccagcg gatccgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     2641 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     2701 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     2761 cttttgcaaa aagctaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     2821 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     2881 ctcatcaatg tatcttatca tgtctggatc cgctgcatta atgaatcggc caacgcgcgg
     2941 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     3001 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     3061 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     3121 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     3181 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     3241 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     3301 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     3361 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     3421 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3481 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     3541 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     3601 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     3661 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     3721 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     3781 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     3841 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     3901 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     3961 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     4021 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     4081 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     4141 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     4201 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     4261 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     4321 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     4381 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     4441 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4501 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     4561 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     4621 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     4681 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     4741 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     4801 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     4861 taggggttcc gcgcacattt ccccgaaaag tgccacctgg gtcgacattg attattgact
     4921 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
     4981 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
     5041 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
     5101 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     5161 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     5221 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     5281 atggtcgagg tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc
     5341 ccaattttgt atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg
     5401 g
//
