LOCUS       sequence-176949-        6694 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Flp recombinase dependent on GFP (Flp-DOG) for rAAV production and 
            expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6694)
  AUTHORS   Tang JC, Drokhlyansky E, Etemad B, Rudolph S, Guo B, Wang S, Ellis 
            EG, Li JZ, Cepko CL
  TITLE     Detection and manipulation of live antigen-expressing cells using 
            conditionally stable nanobodies.
  JOURNAL   Elife. 2016 May 20;5. pii: e15312. doi: 10.7554/eLife.15312.
  PUBMED    27205882
REFERENCE   2  (bases 1 to 6694)
  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: "Elife."; 
            date: "2016-05-20"; volume: "5. pii: e15312. doi"; pages: " 
            10.7554/eLife.15312"
COMMENT     Sequence Label: pAAV-EF1a-Flp-DOG-NW
FEATURES             Location/Qualifiers
     source          1..6694
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   211..351
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   211..340
                     /label=AAV2 ITR
     rep_origin      426..881
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(513..532)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     723..744
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(898..917)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1017..1039
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1077..1095)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1163..1267
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1268..1336,1337..2128)
                     /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     complement(1486..1505)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2299..2887
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2788..2807
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   2949..3078
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        3179..4357
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3410..4348
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     4305..4325
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      4370..4379
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    join(6389..6694,1..171)
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(6588..6607)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
ORIGIN
        1 ttccaggcat gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc
       61 accatattgg ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct
      121 cccaaattgc tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg
      181 taggtaacca cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc
      241 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg
      301 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga
      361 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc
      421 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
      481 gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct
      541 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
      601 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag
      661 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
      721 tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga
      781 tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa
      841 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac
      901 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
      961 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     1021 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     1081 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     1141 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     1201 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     1261 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     1321 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     1381 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     1441 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     1501 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     1561 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     1621 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     1681 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     1741 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     1801 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     1861 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     1921 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     1981 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     2041 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     2101 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     2161 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     2221 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     2281 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2341 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2401 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     2461 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     2521 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     2581 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     2641 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     2701 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     2761 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     2821 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     2881 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     2941 tcacatgtcc tgcaggcagc tgcgcgctcg ctcgctcact gaggccgccc gggcgtcggg
     3001 cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact
     3061 ccatcactag gggttcctgc ggccgcacgc gtaagctttg caaagatgga taaagtttta
     3121 aacagagagg aatctttgca gctaatggac cttctaggtc ttgaaaggag tgggaattgg
     3181 ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg
     3241 aggggtcggc aattgaaccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga
     3301 tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt
     3361 agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg taagtgccgt
     3421 gtgtggttcc cgcgggcctg gcctctttac gggttatggc ccttgcgtgc cttgaattac
     3481 ttccacctgg ctgcagtacg tgattcttga tcccgagctt cgggttggaa gtgggtggga
     3541 gagttcgagg ccttgcgctt aaggagcccc ttcgcctcgt gcttgagttg aggcctggcc
     3601 tgggcgctgg ggccgccgcg tgcgaatctg gtggcacctt cgcgcctgtc tcgctgcttt
     3661 cgataagtct ctagccattt aaaatttttg atgacctgct gcgacgcttt ttttctggca
     3721 agatagtctt gtaaatgcgg gccaagatct gcacactggt atttcggttt ttggggccgc
     3781 gggcggcgac ggggcccgtg cgtcccagcg cacatgttcg gcgaggcggg gcctgcgagc
     3841 gcggccaccg agaatcggac gggggtagtc tcaagctggc cggcctgctc tggtgcctgg
     3901 cctcgcgccg ccgtgtatcg ccccgccctg ggcggcaagg ctggcccggt cggcaccagt
     3961 tgcgtgagcg gaaagatggc cgcttcccgg ccctgctgca gggagctcaa aatggaggac
     4021 gcggcgctcg ggagagcggg cgggtgagtc acccacacaa aggaaaaggg cctttccgtc
     4081 ctcagccgtc gcttcatgtg actccacgga gtaccgggcg ccgtccaggc acctcgatta
     4141 gttctcgagc ttttggagta cgtcgtcttt aggttggggg gaggggtttt atgcgatgga
     4201 gtttccccac actgagtggg tggagactga agttaggcca gcttggcact tgatgtaatt
     4261 ctccttggaa tttgcccttt ttgagtttgg atcttggttc attctcaagc ctcagacagt
     4321 ggttcaaagt ttttttcttc catttcaggt gtcgtgaggt accggatccg ccaccatggc
     4381 cgacgtgcag ctcgtggaat ccggaggagc actggtccag cctgggggat cactgagact
     4441 gtcttgtgtc gcttctggct ttcctgtgaa tcggtactct atgagatggt acaggcaggc
     4501 acctggaaaa gagcgagaat gggtcgccgg aatgtcatct gctggcgata ggagttccta
     4561 cgtggatagc gtgaaaggac ggttcaccat tagaagggac gatgctagaa acaccgtgta
     4621 cctccagatg aactccctga aacctgagga cactgctgtg tactactaca acgtcaatgt
     4681 gggctttgaa tactggggac atggcacaca agtgactgtt agttttgcat gcatggctga
     4741 cgtccaactt gttgagagcg gtggagcact ggtccagcct gggggatcac tgagactgtc
     4801 ttgtgtcgct tctggctttc ctgtgaatcg gtactctatg agatggtaca ggcaggcacc
     4861 tggaaaagag cgagaatggg tcgccggaat gtcatctgct ggcgatagga gttcctacgt
     4921 ggatagcgtg aaaggacggt tcaccattag aagggacgat gctagaaaca ccgtgtacct
     4981 ccagatgaac tccctgaaac ctgaggacac tgctgtgtac tactacaacg tcaatgtggg
     5041 ctttgaatac tggggccatg ggacacaggt tactgttagc ttcgcatgca tgagccagtt
     5101 cgacatcctg tgcaagaccc cccccaaggt gctggtgcgg cagttcgtgg agagattcga
     5161 gaggcccagc ggcgagaaga tcgccagctg tgccgccgag ctgacctacc tgtgctggat
     5221 gatcacccac aacggcaccg ccatcaagag ggccaccttc atgagctaca acaccatcat
     5281 cagcaacagc ctgagcttcg acatcgtgaa caagagcctg cagttcaagt acaagaccca
     5341 gaaggccacc atcctggagg ccagcctgaa gaagctgatc cccgcctggg agttcaccat
     5401 catcccttac aacggccaga agcaccagag cgacatcacc gacatcgtgt ccagcctgca
     5461 gctgcagttc gagagcagcg aggaggccga caagggcaac agccacagca agaagatgct
     5521 gaaggccctg ctgtccgagg gcgagagcat ctgggagatc accgagaaga tcctgaacag
     5581 cttcgagtac accagcaggt tcaccaagac caagaccctg taccagttcc tgttcctggc
     5641 cacattcatc aactgcggca ggttcagcga catcaagaac gtggacccca agagcttcaa
     5701 gctggtgcag aacaagtacc tgggcgtgat cattcagtgc ctggtgaccg agaccaagac
     5761 aagcgtgtcc aggcacatct actttttcag cgccagaggc aggatcgacc ccctggtgta
     5821 cctggacgag ttcctgagga acagcgagcc cgtgctgaag agagtgaaca ggaccggcaa
     5881 cagcagcagc aacaagcagg agtaccagct gctgaaggac aacctggtgc gcagctacaa
     5941 caaggccctg aagaagaacg ccccctaccc catcttcgct atcaagaacg gccctaagag
     6001 ccacatcggc aggcacctga tgaccagctt tctgagcatg aagggcctga ccgagctgac
     6061 aaacgtggtg ggcaactgga gcgacaagag ggcctccgcc gtggccagga ccacctacac
     6121 ccaccagatc accgccatcc ccgaccacta cttcgccctg gtgtccaggt actacgccta
     6181 cgaccccatc agcaaggaga tgatcgccct gaaggacgag accaacccca tcgaggagtg
     6241 gcagcacatc gagcagctga agggcagcgc cgagggcagc atcagatacc ccgcctggaa
     6301 cggcatcatc agccaggagg tgctggacta cctgagcagc tacatcaaca ggcggatcta
     6361 ggaattcgat gctgctcgag agatctacgg gtggcatccc tgtgacccct ccccagtgcc
     6421 tctcctggcc ctggaagttg ccactccagt gcccaccagc cttgtcctaa taaaattaag
     6481 ttgcatcatt ttgtctgact aggtgtcctt ctataatatt atggggtgga ggggggtggt
     6541 atggagcaag gggcaagttg ggaagacaac ctgtagggcc tgcggggtct attgggaacc
     6601 aagctggagt gcagtggcac aatcttggct cactgcaatc tccgcctcct gggttcaagc
     6661 gattctcctg cctcagcctc ccgagttgtt ggga
//
