LOCUS       sequence-469795-        5823 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For Cre-dependent expression of Flp recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5823)
  AUTHORS   Oriol L, Chao M, Kollman GJ, Dowlat DS, Singhal SM, Steinkellner T, 
            Hnasko TS
  TITLE     Ventral tegmental area interneurons revisited: GABA and glutamate 
            projection neurons make local synapses.
  JOURNAL   bioRxiv [Preprint]. 2025 Jan 17:2024.06.07.597996. doi: 
            10.1101/2024.06.07.597996.
  PUBMED    38895464
REFERENCE   2  (bases 1 to 5823)
  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: "bioRxiv 
            [Preprint]. 2025 Jan 17:2024.06.07.597996. doi: 
            10.1101/2024.06.07.597996."
COMMENT     Sequence Label: pAAV-Syn1-DIO-Flpo
FEATURES             Location/Qualifiers
     source          1..5823
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1017..1050
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1061..1094
                     /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)."
     misc_feature    1118..1706
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1171..1191)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1589..1600)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1741..1758)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1747..1954
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   1977..2106
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(2168..2756)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2248..2267)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2927..3718,3719..3787))
                     /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     3550..3569
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3788..3892)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3960..3978
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4016..4038)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4138..4157
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(4174..4629)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4311..4332)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4523..4542
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   4704..4844
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4715..4844
                     /label=AAV2 ITR
     promoter        4880..5327
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(5360..5393)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(5404..5437)
                     /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)."
ORIGIN
        1 aagctggtca tcaggtgcct gccgatgtgg ctcttagggc cgttcttgat agcgaagatg
       61 gggtaggggg cgttcttctt cagggccttg ttgtagctgc gcaccaggtt gtccttcagc
      121 agctggtact cctgcttgtt gctgctgctg ttgccggtcc tgttcactct cttcagcacg
      181 ggctcgctgt tcctcaggaa ctcgtccagg tacaccaggg ggtcgatcct gcctctggcg
      241 ctgaaaaagt agatgtgcct ggacacgctt gtcttggtct cggtcaccag gcactgaatg
      301 atcacgccca ggtacttgtt ctgcaccagc ttgaagctct tggggtccac gttcttgatg
      361 tcgctgaacc tgccgcagtt gatgaatgtg gccaggaaca ggaactggta cagggtcttg
      421 gtcttggtga acctgctggt gtactcgaag ctgttcagga tcttctcggt gatctcccag
      481 atgctctcgc cctcggacag cagggccttc agcatcttct tgctgtggct gttgcccttg
      541 tcggcctcct cgctgctctc gaactgcagc tgcaggctgg acacgatgtc ggtgatgtcg
      601 ctctggtgct tctggccgtt gtaagggatg atggtgaact cccaggcggg gatcagcttc
      661 ttcaggctgg cctccaggat ggtggccttc tgggtcttgt acttgaactg caggctcttg
      721 ttcacgatgt cgaagctcag gctgttgctg atgatggtgt tgtagctcat gaaggtggcc
      781 ctcttgatgg cggtgccgtt gtgggtgatc atccagcaca ggtaggtcag ctcggcggca
      841 cagctggcga tcttctcgcc gctgggcctc tcgaatctct ccacgaactg ccgcaccagc
      901 accttggggg gggtcttgca caggatgtcg aactggctca tggtgggatc cccccaagct
      961 tggaacgtac taaccttgat agctagctag ttggcgcgcc aacctggccg gccttgataa
     1021 cttcgtataa tgtatgctat acgaagttat ccgaatcgca ataacttcgt ataaagtatc
     1081 ctatacgaag ttatcgaaga tcatcaagct tatcgataat caacctctgg attacaaaat
     1141 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     1201 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     1261 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     1321 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     1381 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     1441 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     1501 gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tatgttgcca cctggattct
     1561 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     1621 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     1681 gatctccctt tgggccgcct ccccgcatcg ataccgagcg ctgctcgaga gatcgatctg
     1741 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     1801 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     1861 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     1921 aggattggga agacaatagc aggcatgctg gggacacgtg cggaccgagc ggccgcagga
     1981 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
     2041 gcgaccaaag gtcgcccgac gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc
     2101 tgcaggacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     2161 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     2221 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     2281 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     2341 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     2401 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     2461 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     2521 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     2581 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     2641 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     2701 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     2761 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     2821 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     2881 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     2941 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     3001 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     3061 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     3121 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     3181 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     3241 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     3301 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     3361 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     3421 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     3481 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     3541 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     3601 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     3661 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     3721 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     3781 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     3841 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     3901 cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa
     3961 ataggcgtat cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct
     4021 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     4081 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg
     4141 catcagagca gattgtactg agagtgcacc ataaaattgt aaacgttaat attttgttaa
     4201 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaatagacc gaaatcggca
     4261 aaatccctta taaatcaaaa gaatagcccg agatagagtt gagtgttgtt ccagtttgga
     4321 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     4381 agggcgatgg cccactacgt gaaccatcac ccaaatcaag ttttttgggg tcgaggtgcc
     4441 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     4501 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct aaggcgctgg
     4561 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     4621 agggcgcgta ctatggttgc tttgacgtat gcggtgtgaa ataccgcaca gatgcgtaag
     4681 gagaaaatac cgcatcaggc gcccctgcag gcagctgcgc gctcgctcgc tcactgaggc
     4741 cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg
     4801 agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctgcggcc gcacgcgtct
     4861 gcagagggcc ctgcgtatga gtgcaagtgg gttttaggac caggatgagg cggggtgggg
     4921 gtgcctacct gacgaccgac cccgacccac tggacaagca cccaaccccc attccccaaa
     4981 ttgcgcatcc cctatcagag agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac
     5041 tgccagcttc agcaccgcgg acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc
     5101 gcctcagcac tgaaggcgcg ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg
     5161 gccaccttgg tcgcgtccgc gccgccgccg gcccagccgg accgcaccac gcgaggcgcg
     5221 agataggggg gcacgggcgc gaccatctgc gctgcggcgc cggcgactca gcgctgcctc
     5281 agtctgcggt gggcagcgga ggagtcgtgt cgtgcctgag agcgcagacg cgtggacggt
     5341 acccttcagg tcgacgtcga taacttcgta tagcatacat tatacgaagt tatccgaatc
     5401 gcaataactt cgtataggat actttatacg aagttatttg gcgcgccaaa tttaaataac
     5461 gcaacgtacc ttcgaggggg cccaaggaat tctcagatcc gcctgttgat gtagctgctc
     5521 aggtagtcca gcacctcctg gctgatgatg ccgttccagg cggggtatct gatgctgccc
     5581 tcggcgctgc ccttcagctg ctcgatgtgc tgccactcct cgatggggtt ggtctcgtcc
     5641 ttcagggcga tcatctcctt gctgatgggg tcgtaggcgt agtacctgga caccagggcg
     5701 aagtagtggt cggggatggc ggtgatctgg tgggtgtagg tggtcctggc cacggcggag
     5761 gccctcttgt cgctccagtt gcccaccacg tttgtcagct cggtcaggcc cttcatgctc
     5821 aga
//
