LOCUS       sequence-382820-        7270 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-vector for optogenetic long-term silencing of Cre-positive 
            neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7270)
  AUTHORS   Wiegert Lab
  TITLE     Wiegert Lab Plasmids 2022
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7270)
  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            "Unpublished"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: Ef1a-DIO-Aion-citrine
FEATURES             Location/Qualifiers
     source          1..7270
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1440..1473)
                     /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(1524..1557)
                     /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(1572..2282,2283..2285,2286..2288))
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1614..1635)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1963..1982
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2222..2243
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3224..3233
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3255..3288
                     /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    3339..3372
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3397..3985
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3450..3470)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3868..3879)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4017..4493
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4216..4235)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4533..4673
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4533..4662
                     /label=AAV2 ITR
     rep_origin      4748..5203
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4835..4854)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5045..5066
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5220..5239)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5339..5361
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5399..5417)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5485..5589
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5590..5658,5659..6450)
                     /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(5808..5827)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6621..7209
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7110..7129
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggaa
     1441 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
     1561 gcgagcttct acttgtacag ctcgtccatg ccgagagtga tcccggcggc ggtcacgaac
     1621 tccagcagga ccatgtgatc gcgcttctcg ttggggtctt tgctcagggc ggactggtag
     1681 ctcaggtagt ggttgtcggg cagcagcacg gggccgtcgc cgatgggggt gttctgctgg
     1741 tagtggtcgg cgagctgcac gctgccgtcc tcgatgttgt ggcggatctt gaagttcacc
     1801 ttgatgccgt tcttctgctt gtcggccatg atatagacgt tgtggctgtt gtagttgtac
     1861 tccagcttgt gccccaggat gttgccgtcc tccttgaagt cgatgccctt cagctcgatg
     1921 cggttcacca gggtgtcgcc ctcgaacttc acctcggcgc gggtcttgta gttgccgtcg
     1981 tccttgaaga agatggtgcg ctcctggacg tagccttcgg gcatggcgga cttgaagaag
     2041 tcgtgctgct tcatgtggtc ggggtagcgg gcgaagcaca tcaggccgta gccgaaggtg
     2101 gtcacgaggg tgggccaggg cacgggcagc ttgccggtgg tgcagatgaa cttcagggtc
     2161 agcttgccgt aggtggcatc gccctcgccc tcgccggaca cgctgaactt gtggccgttt
     2221 acgtcgccgt ccagctcgac caggatgggc accaccccgg tgaacagctc ctcgcccttg
     2281 ctcaccatct cgagtgccgc gaccgcgcca gcctcggcct cgtcctccac cagcgtctcg
     2341 acctcaatct cagtgccacc aatgttcaat ttggtggtct tgcgaatgtc gccgtggatg
     2401 aggatatggc tgtggatcag cacgcgcagg tagtggccga gcagacccca gcactgcttc
     2461 gacatcaggt caatgatggt gtggccgacg ttggagccgt accggctcag gacgccgaag
     2521 ccctcggggc cgaggatgaa caggatgggg aacatacccc atgatacgaa gaagagccaa
     2581 gccatgccag tcaccacctg gcgacaccgg cccttcggca cggtgtggta cgcctcaatg
     2641 tagaccttgg ctgcgttgaa gaatgtgtag atgccgtagc acaggcccat caggaagaaa
     2701 atgacacgga cgtatccctt ggacagcgcg gccgtggtgg cccacacgat gccgccgatg
     2761 catgacacca gcagacccat ggtacgcttg ttatagtcgt tcgccagacc cgtaaggttg
     2821 ctcaggtgga taagaatgac aggggcggtc agcagccagc tcgcgtaacg aagccaccgg
     2881 gtcttgttgc cgttggatga gtagatcacc gcaggttcgt caaagctatg gaaatactcg
     2941 atgatgaact tgatcatctg gatcgtggcc acgtaaatgt tctcccagcc gcaagtagac
     3001 ttccaggtct ggtagccgta gaacatcagg cagagcgctg aaagcgcaaa agaaatccac
     3061 tgcagaatgt tggcagccaa cttctcggca tttgttccgt tggatttaag ccaagccaag
     3121 cagaagcact ggccgttgtt cgggatgcaa ataacagaac cattgttctc aagagtgtat
     3181 gaggtttgga agagtccgac ggcagacaaa gcgccgccat agtccatggt ggcgctagca
     3241 gcttgaattc tagcataact tcgtataaag tatcctatac gaagttattt gccttaaccc
     3301 agaaattatc actgttattc tttagaatgg tgcaaagaat aacttcgtat aatgtatgct
     3361 atacgaagtt atgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt
     3421 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     3481 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     3541 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
     3601 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
     3661 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
     3721 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
     3781 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg
     3841 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
     3901 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
     3961 atctcccttt gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt
     4021 ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc
     4081 ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct
     4141 ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct
     4201 gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca
     4261 ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg
     4321 gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt
     4381 tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc
     4441 ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt
     4501 tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac
     4561 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     4621 gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct
     4681 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa
     4741 ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     4801 gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt
     4861 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     4921 cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt
     4981 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     5041 aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt
     5101 gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa
     5161 aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt
     5221 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     5281 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     5341 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     5401 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     5461 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     5521 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     5581 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5641 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5701 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5761 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     5821 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5881 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5941 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     6001 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     6061 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     6121 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     6181 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     6241 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     6301 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     6361 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     6421 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     6481 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6541 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6601 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     6661 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     6721 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     6781 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     6841 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     6901 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     6961 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     7021 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     7081 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     7141 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     7201 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     7261 gctcacatgt
//
