LOCUS       sequence-374767-        6387 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre dependent FlpO expression under the control of CAG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6387)
  TITLE     AAV expression plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6387)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAAV-CAG-Flex-Rev-FlpO
FEATURES             Location/Qualifiers
     source          1..6387
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     14..33
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(252..356)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(382..837)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(519..540)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     731..750
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   909..1038
                     /label=AAV2 ITR
     primer_bind     1903..1925
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1987..2006
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    2041..2074
                     /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)."
     protein_bind    2137..2170
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(2210..3481,3482..3502,3503..3508))
                     /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"
     regulatory      3505..3514
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3560..3593)
                     /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)."
     protein_bind    complement(3664..3697)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3739..4327
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3792..3812)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4210..4221)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4370..4491
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4381..4400)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4435..4454
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4653..4782
                     /label=AAV2 ITR
     rep_origin      complement(5019..5607)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5099..5118)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5778..6387,1..182,183..251))
                     /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"
ORIGIN
        1 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
       61 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      121 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      181 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
      241 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
      301 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
      361 tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
      421 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
      481 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
      541 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
      601 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
      661 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
      721 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
      781 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc
      841 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      901 tacgccagct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg
      961 cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact
     1021 ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc
     1081 catgctctag gaagagtacc attgacgtca ataatgacgt atgttcccat agtaacgcca
     1141 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
     1201 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
     1261 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
     1321 tacgtattag tcatcgctat taccatggtc gaggtgagcc ccacgttctg cttcactctc
     1381 cccatctccc ccccctcccc acccccaatt ttgtatttat ttatttttta attattttgt
     1441 gcagcgatgg gggcgggggg ggggggggsg cgcgccrggg gggggggggg gggggggggg
     1501 ggggggsggg gsgrggsggr gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa
     1561 gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg
     1621 ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg
     1681 cccgccccgg ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc
     1741 tcctccgggc tgtaattagc gcttggttta atgacggctt gtttcttttc tgtggctgcg
     1801 tgaaagcctt gaggggctcc gggagggccc tttgtgcggg gggagcggct cggggctgtc
     1861 cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg
     1921 tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     1981 tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttggatcccc
     2041 ataacttcgt ataaagtatc ctatacgaag ttatatcaaa ataggaagac caatgcttca
     2101 ccatcgaccc gaattgccaa gcatcaccat cgacccataa cttcgtataa tgtatgctat
     2161 acgaagttat actagtgcta gagctagctg atcacttaag gtttagcttt cagatccgcc
     2221 tgttgatgta gctgctcagg tagtccagca cctcctggct gatgatgccg ttccaggcgg
     2281 ggtatctgat gctgccctcg gcgctgccct tcagctgctc gatgtgctgc cactcctcga
     2341 tggggttggt ctcgtccttc agggcgatca tctccttgct gatggggtcg taggcgtagt
     2401 acctggacac cagggcgaag tagtggtcgg ggatggcggt gatctggtgg gtgtaggtgg
     2461 tcctggccac ggcggaggcc ctcttgtcgc tccagttgcc caccacgttt gtcagctcgg
     2521 tcaggccctt catgctcaga aagctggtca tcaggtgcct gccgatgtgg ctcttagggc
     2581 cgttcttgat agcgaagatg gggtaggggg cgttcttctt cagggccttg ttgtagctgc
     2641 gcaccaggtt gtccttcagc agctggtact cctgcttgtt gctgctgctg ttgccggtcc
     2701 tgttcactct cttcagcacg ggctcgctgt tcctcaggaa ctcgtccagg tacaccaggg
     2761 ggtcgatcct gcctctggcg ctgaaaaagt agatgtgcct ggacacgctt gtcttggtct
     2821 cggtcaccag gcactgaatg atcacgccca ggtacttgtt ctgcaccagc ttgaagctct
     2881 tggggtccac gttcttgatg tcgctgaacc tgccgcagtt gatgaatgtg gccaggaaca
     2941 ggaactggta cagggtcttg gtcttggtga acctgctggt gtactcgaag ctgttcagga
     3001 tcttctcggt gatctcccag atgctctcgc cctcggacag cagggccttc agcatcttct
     3061 tgctgtggct gttgcccttg tcggcctcct cgctgctctc gaactgcagc tgcaggctgg
     3121 acacgatgtc ggtgatgtcg ctctggtgct tctggccgtt gtaagggatg atggtgaact
     3181 cccaggcggg gatcagcttc ttcaggctgg cctccaggat ggtggccttc tgggtcttgt
     3241 acttgaactg caggctcttg ttcacgatgt cgaagctcag gctgttgctg atgatggtgt
     3301 tgtagctcat gaaggtggcc ctcttgatgg cggtgccgtt gtgggtgatc atccagcaca
     3361 ggtaggtcag ctcggcggca cagctggcga tcttctcgcc gctgggcctc tcgaatctct
     3421 ccacgaactg ccgcaccagc accttggggg gggtcttgca caggatgtcg aactggctca
     3481 tcaccttcct cttcttctta ggagccatgg tggcggcgga tctcgagtcg cgaacgcgtg
     3541 gcgcgccggt accactagta taacttcgta taggatactt tatacgaagt tatcattggg
     3601 attcttccta ttttgatcca agcatcacca tcgaccctct agtccagatc tcaccatcga
     3661 cccataactt cgtatagcat acattatacg aagttatgtc cctcgaagag gttcgaattc
     3721 gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     3781 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
     3841 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg
     3901 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt
     3961 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact
     4021 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
     4081 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg
     4141 tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc
     4201 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
     4261 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc
     4321 tccccgcatc gataccgtcg acccgggcgg ccgcttcgag cagacatgat aagatacatt
     4381 gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt
     4441 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
     4501 aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagcaag
     4561 taaaacctct acaaatgtgg taaaatcgat aaggatcttc ctagagcatg gctacgtaga
     4621 taagtagcat ggcgggttaa tcattaacta caaggaaccc ctagtgatgg agttggccac
     4681 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     4741 gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcatta atgaatcggc
     4801 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     4861 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4921 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4981 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     5041 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     5101 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5161 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     5221 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     5281 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     5341 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     5401 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     5461 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     5521 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     5581 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5641 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5701 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5761 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5821 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5881 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5941 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     6001 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     6061 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     6121 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     6181 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     6241 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     6301 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     6361 atgcggcgac cgagttgctc ttgcccg
//
