LOCUS       sequence-300336-        7006 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector to drive the expression of dChr2-GFP-P2A-GFP in PV 
            cortical interneuronsunder the control of the E22 regulatory 
            element.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7006)
  AUTHORS   Vormstein-Schneider D, Lin JD, Pelkey KA, Chittajallu R, Guo B, 
            Arias-Garcia MA, Allaway K, Sakopoulos S, Schneider G, Stevenson O, 
            Vergara J, Sharma J, Zhang Q, Franken TP, Smith J, Ibrahim LA, M 
            Astro KJ, Sabri E, Huang S, Favuzzi E, Burbridge T, Xu Q, Guo L, 
            Vogel I, Sanchez V, Saldi GA, Gorissen BL, Yuan X, Zaghloul KA, 
            Devinsky O, Sabatini BL, Batista-Brito R, Reynolds J, Feng G, Fu Z, 
            McBain CJ, Fishell G, Dimidschstein J
  TITLE     Viral manipulation of functionally distinct interneurons in mice, 
            non-human primates and humans.
  JOURNAL   Nat Neurosci. 2020 Dec;23(12):1629-1636. doi: 
            10.1038/s41593-020-0692-9. Epub 2020 Aug 17.
  PUBMED    32807948
REFERENCE   2  (bases 1 to 7006)
  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: 
            "10.1038/s41593-020-0692-9"; journalName: "Nat Neurosci"; date: 
            "2020-12"; volume: "23"; issue: "12"; pages: "1629-1636"
COMMENT     Sequence Label: pAAV-E22-ChR2GFP2x
FEATURES             Location/Qualifiers
     source          1..7006
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          775..907
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     890..909
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      948..957
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             954..1880
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             join(1896..1898,1899..2609)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1896..1898,1899..2609)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             2631..2687
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2700..2720
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(2721..2723,2724..3437)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2721..2723,2724..3434)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     misc_feature    3447..4035
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3500..3520)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3918..3929)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4043..4267
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4269..4409
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4269..4398
                     /label=AAV2 ITR
     rep_origin      4484..4939
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4571..4590)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4781..4802
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4956..4975)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5075..5097
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5135..5153)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5221..5325
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5326..5394,5395..6186)
                     /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(5544..5563)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6357..6945
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6846..6865
                     /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 gctagactgc agagggcccg cggccgcacg cgtatcgatt cggggtttaa
      181 ccgttcagag taattttcca ttataatttt gtgttctaaa taaatagaac cagcttcact
      241 ctagaaacgc ttgtctgagt gctgtacaat gcatgtttaa gtggctctgg tgaaattact
      301 aatcataccc ctgcagtgga taaaatagaa ttgtttgtga gttttgctca ttcgatggga
      361 ggacgctaaa tttataaaag tagactcatg cccttgatag tgaatttaaa taggtgcata
      421 tgtaaacaga tacttactca tttagagaca ccgtcatcag ggagctttgt gtgagaatgt
      481 tttctatgct ccggctcctg ctctgtggta ggggaactaa gcagacatcc caggtccatt
      541 tcctaaccca gtgtctctag ttgagtaatc acacaaggca gacatcccag ctgggactct
      601 atatttaaac ccatcgatgt ttaaacttcc tgcagcccgg gctgggcata aaagtcaggg
      661 cagagccatc tattgcttac atttgcttct agcctgcaag tcgaggagcg cagccttcca
      721 gaagcagagc gcggcgcctt aagctgcaga agttggtcgt gaggcactgg gcacgtaagt
      781 atcaaggtta caagacaggt ttaaggagac caatagaaac tgggcttgtc gagacagaga
      841 agactcttgc gtttctgata ggcacctatt ggtcttactg acatccactt tgcctttctc
      901 tccacaggtg tccactccca gttcaattac agctcttaag aaagctagcc accatggact
      961 atggcggcgc tttgtctgcc gtcggacgcg aacttttgtt cgttactaat cctgtggtgg
     1021 tgaacgggtc cgtcctggtc cctgaggatc aatgttactg tgccggatgg attgaatctc
     1081 gcggcacgaa cggcgctcag accgcgtcaa atgtcctgca gtggcttgca gcaggattca
     1141 gcattttgct gctgatgttc tatgcctacc aaacctggaa atctacatgc ggctgggagg
     1201 agatctatgt gtgcgccatt gaaatggtta aggtgattct cgagttcttt tttgagttta
     1261 agaatccctc tatgctctac cttgccacag gacaccgggt gcagtggctg cgctatgcag
     1321 agtggctgct cacttgtcct gtcatcctta tccgcctgag caacctcacc ggcctgagca
     1381 acgactacag caggagaacc atgggactcc ttgtctcaga catcgggact atcgtgtggg
     1441 gggctaccag cgccatggca accggctatg ttaaagtcat cttcttttgt cttggattgt
     1501 gctatggcgc gaacacattt tttcacgccg ccaaagcata tatcgagggt tatcatactg
     1561 tgccaaaggg tcggtgccgc caggtcgtga ccggcatggc atggctgttt ttcgtgagct
     1621 ggggtatgtt cccaattctc ttcattttgg ggcccgaagg ttttggcgtc ctgagcgtct
     1681 atggctccac cgtaggtcac acgattattg atctgatgag taaaaattgt tgggggttgt
     1741 tgggacacta cctgcgcgtc ctgatccacg agcacatatt gattcacgga gatatccgca
     1801 aaaccaccaa actgaacatc ggcggaacgg agatcgaggt cgagactctc gtcgaagacg
     1861 aagccgaggc cggagccgtg ccagctgcag ccacagtgag caagggcgag gagctgttca
     1921 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     1981 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     2041 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     2101 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     2161 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     2221 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     2281 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     2341 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     2401 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     2461 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     2521 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     2581 tcactctcgg catggacgag ctgtacaagt ccggagataa cggaagcgga gccactaact
     2641 tctccctgtt gaaacaagca ggggatgtcg aagagaatcc cgggccatct agaatggccc
     2701 ccaagaagaa gaggaaggtg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca
     2761 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     2821 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     2881 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     2941 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     3001 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     3061 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     3121 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     3181 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     3241 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     3301 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     3361 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     3421 acgagctgta caagtaatgg cgcgccaatc aacctctgga ttacaaaatt tgtgaaagat
     3481 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     3541 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     3601 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     3661 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     3721 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     3781 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     3841 agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt
     3901 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     3961 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     4021 gggccgcctc cccgcctgga tcctgtgcct tctagttgcc agccatctgt tgtttgcccc
     4081 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     4141 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     4201 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     4261 tctatggcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     4321 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     4381 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     4441 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     4501 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     4561 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     4621 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     4681 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     4741 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     4801 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     4861 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     4921 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     4981 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     5041 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     5101 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     5161 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     5221 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     5281 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     5341 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     5401 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     5461 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     5521 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     5581 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     5641 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     5701 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     5761 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     5821 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     5881 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     5941 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     6001 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     6061 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     6121 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     6181 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     6241 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     6301 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     6361 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     6421 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     6481 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     6541 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     6601 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     6661 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     6721 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     6781 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     6841 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     6901 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     6961 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
