LOCUS       sequence-170322-        6892 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An AAV vector expresses FLPo in a Cre recombinase-dependent manner..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6892)
  AUTHORS   Zingg B, Chou XL, Zhang ZG, Mesik L, Liang F, Tao HW, Zhang LI
  TITLE     AAV-Mediated Anterograde Transsynaptic Tagging: Mapping 
            Corticocollicular Input-Defined Neural Pathways for Defense 
            Behaviors.
  JOURNAL   Neuron. 2017 Jan 4;93(1):33-47. doi: 10.1016/j.neuron.2016.11.045. 
            Epub 2016 Dec 15.
  PUBMED    27989459
REFERENCE   2  (bases 1 to 6892)
  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"; doi: 
            "10.1016/j.neuron.2016.11.045"; journalName: "Neuron"; date: 
            "2017-01-4- 4"; volume: "93"; issue: "1"; pages: "33-47"
COMMENT     Sequence Label: AAV pEF1a-DIO-FLPo-WPRE-hGHpA
FEATURES             Location/Qualifiers
     source          1..6892
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   13..142
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        243..1421
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          474..1412
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1369..1389
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1452..1485)
                     /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(1536..1569)
                     /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(1581..2852,2853..2873,2874..2879))
                     /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"
     protein_bind    2889..2922
                     /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    2973..3006
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3031..3619
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3084..3104)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3502..3513)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3651..4127
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3850..3869)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4167..4307
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4167..4296
                     /label=AAV2 ITR
     rep_origin      4382..4837
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4469..4488)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4679..4700
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4854..4873)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4973..4995
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5033..5051)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5119..5223
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5224..5292,5293..6084)
                     /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(5442..5461)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6255..6843
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6744..6763
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttgctcacat gtcctgcagg cagctgcgcg ctcgctcgct cactgaggcc gcccgggcgt
       61 cgggcgacct ttggtcgccc ggcctcagtg agcgagcgag cgcgcagaga gggagtggcc
      121 aactccatca ctaggggttc ctgcggccgc acgcgtaagc tttgcaaaga tggataaagt
      181 tttaaacaga gaggaatctt tgcagctaat ggaccttcta ggtcttgaaa ggagtgggaa
      241 ttggctccgg tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg
      301 ggggaggggt cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa
      361 gtgatgtcgt gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg
      421 cagtagtcgc cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtaagtg
      481 ccgtgtgtgg ttcccgcggg cctggcctct ttacgggtta tggcccttgc gtgccttgaa
      541 ttacttccac ctggctgcag tacgtgattc ttgatcccga gcttcgggtt ggaagtgggt
      601 gggagagttc gaggccttgc gcttaaggag ccccttcgcc tcgtgcttga gttgaggcct
      661 ggcctgggcg ctggggccgc cgcgtgcgaa tctggtggca ccttcgcgcc tgtctcgctg
      721 ctttcgataa gtctctagcc atttaaaatt tttgatgacc tgctgcgacg ctttttttct
      781 ggcaagatag tcttgtaaat gcgggccaag atctgcacac tggtatttcg gtttttgggg
      841 ccgcgggcgg cgacggggcc cgtgcgtccc agcgcacatg ttcggcgagg cggggcctgc
      901 gagcgcggcc accgagaatc ggacgggggt agtctcaagc tggccggcct gctctggtgc
      961 ctggcctcgc gccgccgtgt atcgccccgc cctgggcggc aaggctggcc cggtcggcac
     1021 cagttgcgtg agcggaaaga tggccgcttc ccggccctgc tgcagggagc tcaaaatgga
     1081 ggacgcggcg ctcgggagag cgggcgggtg agtcacccac acaaaggaaa agggcctttc
     1141 cgtcctcagc cgtcgcttca tgtgactcca cggagtaccg ggcgccgtcc aggcacctcg
     1201 attagttctc gagcttttgg agtacgtcgt ctttaggttg gggggagggg ttttatgcga
     1261 tggagtttcc ccacactgag tgggtggaga ctgaagttag gccagcttgg cacttgatgt
     1321 aattctcctt ggaatttgcc ctttttgagt ttggatcttg gttcattctc aagcctcaga
     1381 cagtggttca aagttttttt cttccatttc aggtgtcgtg aggtaccgga tcctctagag
     1441 tcgactccgg aataacttcg tataggatac tttatacgaa gttatgcaga atggtagctg
     1501 gattgtagct gctattagca atatgaaacc tcttaataac ttcgtatagc atacattata
     1561 cgaagttatg gcgcgcctta tcagatccgc ctgttgatgt agctgctcag gtagtccagc
     1621 acctcctggc tgatgatgcc gttccaggcg gggtatctga tgctgccctc ggcgctgccc
     1681 ttcagctgct cgatgtgctg ccactcctcg atggggttgg tctcgtcctt cagggcgatc
     1741 atctccttgc tgatggggtc gtaggcgtag tacctggaca ccagggcgaa gtagtggtcg
     1801 gggatggcgg tgatctggtg ggtgtaggtg gtcctggcca cggcggaggc cctcttgtcg
     1861 ctccagttgc ccaccacgtt tgtcagctcg gtcaggccct tcatgctcag aaagctggtc
     1921 atcaggtgcc tgccgatgtg gctcttaggg ccgttcttga tagcgaagat ggggtagggg
     1981 gcgttcttct tcagggcctt gttgtagctg cgcaccaggt tgtccttcag cagctggtac
     2041 tcctgcttgt tgctgctgct gttgccggtc ctgttcactc tcttcagcac gggctcgctg
     2101 ttcctcagga actcgtccag gtacaccagg gggtcgatcc tgcctctggc gctgaaaaag
     2161 tagatgtgcc tggacacgct tgtcttggtc tcggtcacca ggcactgaat gatcacgccc
     2221 aggtacttgt tctgcaccag cttgaagctc ttggggtcca cgttcttgat gtcgctgaac
     2281 ctgccgcagt tgatgaatgt ggccaggaac aggaactggt acagggtctt ggtcttggtg
     2341 aacctgctgg tgtactcgaa gctgttcagg atcttctcgg tgatctccca gatgctctcg
     2401 ccctcggaca gcagggcctt cagcatcttc ttgctgtggc tgttgccctt gtcggcctcc
     2461 tcgctgctct cgaactgcag ctgcaggctg gacacgatgt cggtgatgtc gctctggtgc
     2521 ttctggccgt tgtaagggat gatggtgaac tcccaggcgg ggatcagctt cttcaggctg
     2581 gcctccagga tggtggcctt ctgggtcttg tacttgaact gcaggctctt gttcacgatg
     2641 tcgaagctca ggctgttgct gatgatggtg ttgtagctca tgaaggtggc cctcttgatg
     2701 gcggtgccgt tgtgggtgat catccagcac aggtaggtca gctcggcggc acagctggcg
     2761 atcttctcgc cgctgggcct ctcgaatctc tccacgaact gccgcaccag caccttgggg
     2821 ggggtcttgc acaggatgtc gaactggctc atcaccttcc tcttcttctt aggagccatg
     2881 gtgctagcat aacttcgtat aaagtatcct atacgaagtt atttgcctta acccagaaat
     2941 tatcactgtt attctttaga atggtgcaaa gaataacttc gtataatgta tgctatacga
     3001 agttatgaat tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa
     3061 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     3121 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     3181 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     3241 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     3301 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     3361 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     3421 gggaaatcat cgtcctttcc ttggctgctc gcctatgttg ccacctggat tctgcgcggg
     3481 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     3541 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     3601 ctttgggccg cctccccgca tcgataccga gcgctgctcg agagatctac gggtggcatc
     3661 cctgtgaccc ctccccagtg cctctcctgg ccctggaagt tgccactcca gtgcccacca
     3721 gccttgtcct aataaaatta agttgcatca ttttgtctga ctaggtgtcc ttctataata
     3781 ttatggggtg gaggggggtg gtatggagca aggggcaagt tgggaagaca acctgtaggg
     3841 cctgcggggt ctattgggaa ccaagctgga gtgcagtggc acaatcttgg ctcactgcaa
     3901 tctccgcctc ctgggttcaa gcgattctcc tgcctcagcc tcccgagttg ttgggattcc
     3961 aggcatgcat gaccaggctc agctaatttt tgtttttttg gtagagacgg ggtttcacca
     4021 tattggccag gctggtctcc aactcctaat ctcaggtgat ctacccacct tggcctccca
     4081 aattgctggg attacaggcg tgaaccactg ctcccttccc tgtccttctg attttgtagg
     4141 taaccacgtg cggaccgagc ggccgcagga acccctagtg atggagttgg ccactccctc
     4201 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt
     4261 tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg
     4321 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag
     4381 tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4441 gctacacttg ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4501 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4561 agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg
     4621 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     4681 ggactcttgt tccaaactgg aacaacactc aactctatct cgggctattc ttttgattta
     4741 taagggattt tgccgatttc ggtctattgg ttaaaaaatg agctgattta acaaaaattt
     4801 aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc
     4861 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     4921 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     4981 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
     5041 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     5101 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     5161 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     5221 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     5281 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     5341 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     5401 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     5461 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     5521 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     5581 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     5641 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     5701 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     5761 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     5821 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     5881 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     5941 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     6001 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     6061 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     6121 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     6181 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     6241 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     6301 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     6361 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     6421 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     6481 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     6541 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     6601 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     6661 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     6721 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     6781 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     6841 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc tt
//
