LOCUS       sequence-276175-        7427 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for Cre-dependent expression of Venus-tagged parapinopsin
            driven by the Ef1a promoter. Used for photoswitchable control of 
            inhibitory GPCR signaling cascades..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7427)
  AUTHORS   Copits BA, Gowrishankar R, O'Neill PR, Li JN, Girven KS, Yoo JJ, 
            Meshik X, Parker KE, Spangler SM, Elerding AJ, Brown BJ, Shirley SE,
            Ma KKL, Vasquez AM, Stander MC, Kalyanaraman V, Vogt SK, Samineni 
            VK, Patriarchi T, Tian L, Gautam N, Sunahara RK, Gereau RW 4th, 
            Bruchas MR
  TITLE     A photoswitchable GPCR-based opsin for presynaptic inhibition.
  JOURNAL   Neuron. 2021 Jun 2;109(11):1791-1809.e11. doi: 
            10.1016/j.neuron.2021.04.026. Epub 2021 May 11.
  PUBMED    33979635
REFERENCE   2  (bases 1 to 7427)
  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.2021.04.026"; journalName: "Neuron"; date: 
            "2021-06-2- 2"; volume: "109"; issue: "11"; pages: "1791-1809.e11"
COMMENT     Sequence Label: pAAV-Ef1a-DIO-PPO-Venus
FEATURES             Location/Qualifiers
     source          1..7427
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        61..1239
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          292..1230
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1187..1207
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1270..1303)
                     /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(1354..1387)
                     /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(1421..2134,2135..2137,2138..2140))
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1466..1487)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1815..1834
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2074..2095
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2137..2146
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3242..3275
                     /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    3326..3359
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3384..3972
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3437..3457)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3855..3866)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4004..4480
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4203..4222)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4520..4660
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4520..4649
                     /label=AAV2 ITR
     rep_origin      4735..5190
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4822..4841)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5032..5053
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5207..5226)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5326..5348
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5386..5404)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5472..5576
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5577..5645,5646..6437)
                     /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(5795..5814)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6608..7196
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7097..7116
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   7258..7387
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 taaacagaga ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt
       61 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
      121 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      181 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      241 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      301 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      361 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      421 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      481 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      541 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      601 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      661 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      721 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      781 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      841 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      901 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
      961 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1021 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1081 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1141 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1201 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc
     1261 gactccggaa taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga
     1321 ttgtagctgc tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg
     1381 aagttatggc gcgccagcgg gtttaaacgg gccctctaga ttacttgtac agctcgtcca
     1441 tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga tcgcgcttct
     1501 cgttggggtc tttgctcagt ttggactggt agctcaggta gtggttgtcg ggcagcagca
     1561 cggggccgtc gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc acgccgccgt
     1621 cctcgatgtt gtggcggatc ttgaagttgg ccttgatgcc gttcttctgc ttgtcggcgg
     1681 tgatatagac gttgtggctg ttgtagttgt actccagctt gtgccccagg atgttgccgt
     1741 cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact
     1801 tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg cgctcctgga
     1861 cgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc
     1921 gggcgaagca ctgcaggccg tagcccaggg tggtcacgag ggtgggccag ggcacgggca
     1981 gcttgccggt ggtgcagatc agcttcaggg tcagcttgcc gtaggtggca tcgccctcgc
     2041 cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg
     2101 gcaccacccc ggtgaacagc tcctcgccct tgctcaccat ggcggccgcg ctcggggaga
     2161 cctgccccgg cgcggacgcc agggtcacgc tggtggccga cgtggaggcc gccgtcgccg
     2221 actccgagga aggctctgcc caggggttgc gtccgcagag caggaagggc acggcgcagt
     2281 cccggaactg gcggttcatg aagatgtaga tgatggggtt gtagaccgtg ctcgtcttgg
     2341 tcaggtacat gggcagtgtg gcgatgaccg ggtcgatgta cacgtcgggc ttggtcacca
     2401 cgatcatggc gaagagcgcg tagggcagcc agcagacgag gaaggccacc accatgacca
     2461 ccaccatgcg cgacacctgc gcctccgcct tggcggtgct gccgctctcg cccatcccca
     2521 gcttggccac ctggtggagg gtccacatga ggcggccgta cgccaccacg atgacgagga
     2581 aggggatggc gaagcagaag gcgaagtagc tcgtaatgta cgacacgttg gcggggtcgc
     2641 ggctgtacca gtcgggcgcg cacgacgtcc gcacgccctc cagctcgtag ctgccccagc
     2701 cgaagagcgg cggcgtgttc cacacgaacg accagagcca ggcccaggcg atgcccagca
     2761 gcgcgtggcg ccgcgtgaac atcagcgtgc ccagcggctt gcacaccacc acgaagcgat
     2821 cgacggcgat cacggcgatc gtgcacagag cggcgatgcc gaagaaagcg acggcgaacc
     2881 cttcgatgac gcagccgacg cggccgaggt tgaagtaccc gacggcgttg gtctccacga
     2941 cgaggctggc gccgaacacc gtgacgccca ggtccgccac ggcgaggttg acgagcgaga
     3001 agttgagcgg gtggcgtagc tggcggtggc gcagggtgac gatgacgacg gtgctgttga
     3061 gcacgagcga ggcgatggtg aacacggcca tgatcacggc gaggatggtg aagccgtagc
     3121 ggggcatcaa cgggacctcg ccgttgggca ggaggtcgag cgaggtcaag ttctccatga
     3181 attccaccac actggactag tggatccgag ctcggtacca agcttaagtt taaacgctag
     3241 cataacttcg tataaagtat cctatacgaa gttatttgcc ttaacccaga aattatcact
     3301 gttattcttt agaatggtgc aaagaataac ttcgtataat gtatgctata cgaagttatg
     3361 aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt gaaagattga
     3421 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt
     3481 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt
     3541 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg
     3601 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg
     3661 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc
     3721 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat
     3781 catcgtcctt tccttggctg ctcgcctatg ttgccacctg gattctgcgc gggacgtcct
     3841 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg
     3901 ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg
     3961 ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc tacgggtggc atccctgtga
     4021 cccctcccca gtgcctctcc tggccctgga agttgccact ccagtgccca ccagccttgt
     4081 cctaataaaa ttaagttgca tcattttgtc tgactaggtg tccttctata atattatggg
     4141 gtggaggggg gtggtatgga gcaaggggca agttgggaag acaacctgta gggcctgcgg
     4201 ggtctattgg gaaccaagct ggagtgcagt ggcacaatct tggctcactg caatctccgc
     4261 ctcctgggtt caagcgattc tcctgcctca gcctcccgag ttgttgggat tccaggcatg
     4321 catgaccagg ctcagctaat ttttgttttt ttggtagaga cggggtttca ccatattggc
     4381 caggctggtc tccaactcct aatctcaggt gatctaccca ccttggcctc ccaaattgct
     4441 gggattacag gcgtgaacca ctgctccctt ccctgtcctt ctgattttgt aggtaaccac
     4501 gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc ctctctgcgc
     4561 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg
     4621 gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt
     4681 ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc
     4741 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     4801 ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     4861 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     4921 tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc
     4981 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     5041 tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat ttataaggga
     5101 ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     5161 attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg
     5221 atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     5281 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     5341 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
     5401 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     5461 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     5521 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     5581 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     5641 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     5701 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5761 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     5821 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5881 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     5941 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     6001 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     6061 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     6121 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     6181 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6241 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6301 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6361 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6421 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     6481 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     6541 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     6601 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     6661 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     6721 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     6781 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     6841 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     6901 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     6961 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     7021 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     7081 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     7141 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     7201 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgtcct
     7261 gcaggcagct gcgcgctcgc tcgctcactg aggccgcccg ggcgtcgggc gacctttggt
     7321 cgcccggcct cagtgagcga gcgagcgcgc agagagggag tggccaactc catcactagg
     7381 ggttcctgcg gccgcacgcg taagctttgc aaagatggat aaagttt
//
