LOCUS       sequence-177045-        5348 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses FlpO under pCAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5348)
  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 5348)
  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: pCAG-FlpO
FEATURES             Location/Qualifiers
     source          1..5348
                     /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"
     regulatory      500..509
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(506..511,512..532,533..1804)
                     /codon_start=1
                     /product="nuclear-targeted site-specific recombinase"
                     /label=FLPo
                     /note="FLPo is a mouse codon-optimized fusion between the 
                     SV40 nuclear localization signal and a thermostable version
                     of the Saccharomyces cerevisiae site-specific recombinase 
                     FLP.  Recombination occurs at FRT sequences."
                     /translation="MAPKKKRKVMSQFDILCKTPPKVLVRQFVERFERPSGEKIASCAA
                     ELTYLCWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKK
                     LIPAWEFTIIPYNGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESI
                     WEITEKILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGV
                     IIQCLVTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEY
                     QLLKDNLVRSYNKALKKNAPYPIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWS
                     DKRASAVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQL
                     KGSAEGSIRYPAWNGIISQEVLDYLSSYINRRI"
     primer_bind     complement(1890..1909)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    1955..2010
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2009..2028)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2371..2387)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2371..2387)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2384..2406)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2395..2411
                     /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(2419..2425,2426..2443,2444..2449))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2464..2485
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      2570..2704
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2724..2858
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2761..2780)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2815..2834
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2926..2943)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3097..3685)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3177..3196)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3856..4647,4648..4716))
                     /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     4479..4498
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4717..4821)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4852..5231
                     /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 acgcgtgccg ccaccatggc tcctaagaag aagaggaagg tgatgagcca
      541 gttcgacatc ctgtgcaaga ccccccccaa ggtgctggtg cggcagttcg tggagagatt
      601 cgagaggccc agcggcgaga agatcgccag ctgtgccgcc gagctgacct acctgtgctg
      661 gatgatcacc cacaacggca ccgccatcaa gagggccacc ttcatgagct acaacaccat
      721 catcagcaac agcctgagct tcgacatcgt gaacaagagc ctgcagttca agtacaagac
      781 ccagaaggcc accatcctgg aggccagcct gaagaagctg atccccgcct gggagttcac
      841 catcatccct tacaacggcc agaagcacca gagcgacatc accgacatcg tgtccagcct
      901 gcagctgcag ttcgagagca gcgaggaggc cgacaagggc aacagccaca gcaagaagat
      961 gctgaaggcc ctgctgtccg agggcgagag catctgggag atcaccgaga agatcctgaa
     1021 cagcttcgag tacaccagca ggttcaccaa gaccaagacc ctgtaccagt tcctgttcct
     1081 ggccacattc atcaactgcg gcaggttcag cgacatcaag aacgtggacc ccaagagctt
     1141 caagctggtg cagaacaagt acctgggcgt gatcattcag tgcctggtga ccgagaccaa
     1201 gacaagcgtg tccaggcaca tctacttttt cagcgccaga ggcaggatcg accccctggt
     1261 gtacctggac gagttcctga ggaacagcga gcccgtgctg aagagagtga acaggaccgg
     1321 caacagcagc agcaacaagc aggagtacca gctgctgaag gacaacctgg tgcgcagcta
     1381 caacaaggcc ctgaagaaga acgcccccta ccccatcttc gctatcaaga acggccctaa
     1441 gagccacatc ggcaggcacc tgatgaccag ctttctgagc atgaagggcc tgaccgagct
     1501 gacaaacgtg gtgggcaact ggagcgacaa gagggcctcc gccgtggcca ggaccaccta
     1561 cacccaccag atcaccgcca tccccgacca ctacttcgcc ctggtgtcca ggtactacgc
     1621 ctacgacccc atcagcaagg agatgatcgc cctgaaggac gagaccaacc ccatcgagga
     1681 gtggcagcac atcgagcagc tgaagggcag cgccgagggc agcatcagat accccgcctg
     1741 gaacggcatc atcagccagg aggtgctgga ctacctgagc agctacatca acaggcggat
     1801 ctgagcggcc gcactcctca ggtgcaggct gcctatcaga aggtggtggc tggtgtggcc
     1861 aatgccctgg ctcacaaata ccactgagat ctttttccct ctgccaaaaa ttatggggac
     1921 atcatgaagc cccttgagca tctgacttct ggctaataaa ggaaatttat tttcattgca
     1981 atagtgtgtt ggaatttttt gtgtctctca ctcggaagga catatgggag ggcaaatcat
     2041 ttaaaacatc agaatgagta tttggtttag agtttggcaa catatgccca tatgctggct
     2101 gccatgaaca aaggttggct ataaagaggt catcagtata tgaaacagcc ccctgctgtc
     2161 cattccttat tccatagaaa agccttgact tgaggttaga ttttttttat attttgtttt
     2221 gtgttatttt tttctttaac atccctaaaa ttttccttac atgttttact agccagattt
     2281 ttcctcctct cctgactact cccagtcata gctgtccctc ttctcttatg gagatccctc
     2341 gacctgcagc ccaagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     2401 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     2461 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     2521 aaacctgtcg tgccagcgga tccgcatctc aattagtcag caaccatagt cccgccccta
     2581 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     2641 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     2701 ggcctaggct tttgcaaaaa gctaacttgt ttattgcagc ttataatggt tacaaataaa
     2761 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     2821 tgtccaaact catcaatgta tcttatcatg tctggatccg ctgcattaat gaatcggcca
     2881 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     2941 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     3001 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     3061 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     3121 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     3181 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     3241 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     3301 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     3361 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     3421 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     3481 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     3541 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     3601 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     3661 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     3721 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     3781 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     3841 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     3901 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     3961 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     4021 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     4081 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     4141 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     4201 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     4261 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     4321 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     4381 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     4441 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     4501 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     4561 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     4621 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     4681 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     4741 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     4801 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgggt cgacattgat
     4861 tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
     4921 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
     4981 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
     5041 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
     5101 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
     5161 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
     5221 ctattaccat ggtcgaggtg agccccacgt tctgcttcac tctccccatc tcccccccct
     5281 ccccaccccc aattttgtat ttatttattt tttaattatt ttgtgcagcg atgggggcgg
     5341 gggggggg
//
