LOCUS       sequence-421862-        6646 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV construct expressing mCherry-tagged channelrhodopsin-2 driven by
            cholinergic neuron-targeting enhancer. Alias: 
            pAAV_S9E27_ChR2_mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6646)
  AUTHORS   Furlanis E, Dai M, Leyva Garcia B, Tran T, Vergara J, Pereira A, 
            Gorissen BL, Wills S, Vlachos A, Hairston A, Dwivedi D, Du S, 
            McMahon J, Huang S, Morabito A, Vazquez A, Kim S, Lee AT, Chang EF, 
            Razzaq T, Qazi A, Vargish G, Yuan X, Caccavano A, Hunt S, 
            Chittajallu R, McLean N, Hewitt L, Paranzino E, Rice H, Cummins AC, 
            Plotnikova A, Mohanty A, Tangen AC, Shin JH, Azadi R, Eldridge MAG, 
            Alvarez VA, Averbeck BB, Alyahyay M, Vallejo TR, Soheib M, Vattino 
            LG, MacGregor CP, Chatain CP, Banks E, Olah VJ, Naskar S, Hill S, 
            Liebergall S, Badiani R, Hyde L, Hanley E, Xu Q, Allaway KC, 
            Goldberg EM, Rowan MJM, Nowakowski TJ, Lee S, Favuzzi E, Kaeser PS, 
            Sjulson L, Batista-Brito R, Takesian AE, Ibrahim LA, Iqbal A, Pelkey
            KA, McBain CJ, Dimidschstein J, Fishell G, Wang Y
  TITLE     An enhancer-AAV toolbox to target and manipulate distinct 
            interneuron subtypes.
  JOURNAL   Neuron. 2025 May 21;113(10):1525-1547.e15. doi: 
            10.1016/j.neuron.2025.05.002.
  PUBMED    40403705
REFERENCE   2  (bases 1 to 6646)
  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.2025.05"; journalName: "Neuron"; date: 
            "2025-05-21- 21"; volume: "113"; issue: "10"; pages: "1525-1547.e15"
COMMENT     Sequence Label: pAAV_BiCHATe27_ChR2_mCherry
FEATURES             Location/Qualifiers
     source          1..6646
                     /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          1150..1282
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1297..1306
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1303..2229
                     /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             2242..2949
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2384..2402)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2652..2671
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    2992..3580
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3045..3065)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3463..3474)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3749..3830
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3786..3805)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3840..3859
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   3909..4049
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3909..4038
                     /label=AAV2 ITR
     rep_origin      4124..4579
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4211..4230)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4421..4442
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4596..4615)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4715..4737
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4775..4793)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4861..4965
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4966..5034,5035..5826)
                     /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(5184..5203)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5997..6585
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6486..6505
                     /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 gcgtatcgat cctcgggtaa tttgtaaaga caatttttct
      181 tttccccctc tgaacctagg ccttaggctg gctaggcaag tctccactac agccccgccc
      241 actccatctc ggctgtggct cgcgggaaag gcagccccct acagttagtc tccccagcca
      301 tcttctccgg ttcccctgat gctctcatgg ccccagccat cctagagctg gtgcgccatc
      361 gctcaccttt agcattcggc ctgctggggg tgagcttttg ggttagaagg aacaaggcat
      421 cagatggggt cgtgtttttt tacattccaa tcagttttcg gaggggaaga acagacattt
      481 ttgtcagcta attgcattta attacagaga caatcgctgt ttccgaacgt tgttgtgtcc
      541 actccctcat ccttcaatca gttcaagcac atccggcccc agctgggagc cttttaattg
      601 cataatttgg atttcacttc tggaaagaaa cttgagaaag actccgtgag ggtctccatt
      661 aacacaatca gctccccagg cacccggggc tggccctaat tgacccatct cagctaatgc
      721 ctctttccaa agaacgcccg cgtcatttct ttggcttgtt tgtgctattg tatcctgttt
      781 tagtgtcttg atcttaaaca tctttaaaga gggcactcag tggcttgttt aactggaagt
      841 ggtccagctg gtttggaagg acagtgcatc aggtgatgta agtccaaggg taggaatttc
      901 tggcctctcc ttcccctgcc cattgcaccg ctctccccaa caggaagtag ctacacatga
      961 taaggtgcac tgtggcttct aggacttatc gatgtttaaa cttcctgcag cccgggctgg
     1021 gcataaaagt cagggcagag ccatctattg cttacatttg cttctagcct gcaagtcgag
     1081 gagcgcagcc ttccagaagc agagcgcggc gccttaagct gcagaagttg gtcgtgaggc
     1141 actgggcacg taagtatcaa ggttacaaga caggtttaag gagaccaata gaaactgggc
     1201 ttgtcgagac agagaagact cttgcgtttc tgataggcac ctattggtct tactgacatc
     1261 cactttgcct ttctctccac agcgagctcg ctagtagcca ccatggacta tggcggcgct
     1321 ttgtctgccg tcggacgcga acttttgttc gttactaatc ctgtggtggt gaacgggtcc
     1381 gtcctggtcc ctgaggatca atgttactgt gccggatgga ttgaatctcg cggcacgaac
     1441 ggcgctcaga ccgcgtcaaa tgtcctgcag tggcttgcag caggattcag cattttgctg
     1501 ctgatgttct atgcctacca aacctggaaa tctacatgcg gctgggagga gatctatgtg
     1561 tgcgccattg aaatggttaa ggtgattctc gagttctttt ttgagtttaa gaatccctct
     1621 atgctctacc ttgccacagg acaccgggtg cagtggctgc gctatgcaga gtggctgctc
     1681 acttgtcctg tcatccttat ccgcctgagc aacctcaccg gcctgagcaa cgactacagc
     1741 aggagaacca tgggactcct tgtctcagac atcgggacta tcgtgtgggg ggctaccagc
     1801 gccatggcaa ccggctatgt taaagtcatc ttcttttgtc ttggattgtg ctatggcgcg
     1861 aacacatttt ttcacgccgc caaagcatat atcgagggtt atcatactgt gccaaagggt
     1921 cggtgccgcc aggtcgtgac cggcatggca tggctgtttt tcgtgagctg gggtatgttc
     1981 ccaattctct tcattttggg gcccgaaggt tttggcgtcc tgagcgtcta tggctccacc
     2041 gtaggtcaca cgattattga tctgatgagt aaaaattgtt gggggttgtt gggacactac
     2101 ctgcgcgtcc tgatccacga gcacatattg attcacggag atatccgcaa aaccaccaaa
     2161 ctgaacatcg gcggaacgga gatcgaggtc gagactctcg tcgaagacga agccgaggcc
     2221 ggagccgtgc cagcggccgc cgtgagcaag ggcgaggagg ataacatggc catcatcaag
     2281 gagttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc
     2341 gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc
     2401 aagggtggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc
     2461 aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag
     2521 ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag
     2581 gactcctccc tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc
     2641 ccctccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc ctccgagcgg
     2701 atgtaccccg aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac
     2761 ggcggccact acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg
     2821 cccggcgcct acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc
     2881 atcgtggaac agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg
     2941 tacaagtaaa acgcgtgccc ctgcagggaa ttcgatatca agcttatcga taatcaacct
     3001 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     3061 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     3121 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     3181 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
     3241 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
     3301 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     3361 gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctatgtt
     3421 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
     3481 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
     3541 cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg agcgctgctc
     3601 gagagatctg tgatagcggc catcaagctg gccgcgactc tagatcataa tcagccatac
     3661 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     3721 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     3781 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     3841 tggtttgtcc aaactcatca atgtatcagc ttatcgatac cgcatgcacg tgcggaccga
     3901 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     3961 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     4021 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     4081 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     4141 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     4201 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     4261 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     4321 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     4381 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     4441 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     4501 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     4561 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     4621 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     4681 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     4741 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     4801 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     4861 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     4921 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     4981 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     5041 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     5101 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     5161 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     5221 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     5281 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     5341 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     5401 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     5461 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     5521 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     5581 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     5641 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     5701 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     5761 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     5821 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     5881 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     5941 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     6001 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     6061 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     6121 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     6181 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     6241 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     6301 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     6361 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     6421 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     6481 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     6541 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     6601 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
