LOCUS       sequence-236122-        6533 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express the genetically-encoded fluorescent acethycholine(ACh) 
            sensor GACh2.0.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6533)
  AUTHORS   Jing M, Zhang P, Wang G, Feng J, Mesik L, Zeng J, Jiang H, Wang S, 
            Looby JC, Guagliardo NA, Langma LW, Lu J, Zuo Y, Talmage DA, Role 
            LW, Barrett PQ, Zhang LI, Luo M, Song Y, Zhu JJ, Li Y
  TITLE     A genetically encoded fluorescent acetylcholine indicator for in 
            vitro and in vivo studies.
  JOURNAL   Nat Biotechnol. 2018 Sep;36(8):726-737. doi: 10.1038/nbt.4184. Epub 
            2018 Jul 9.
  PUBMED    29985477
REFERENCE   2  (bases 1 to 6533)
  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/nbt.4184"; 
            journalName: "Nat Biotechnol"; date: "2018-09"; volume: "36"; issue:
            "8"; pages: "726-737"
COMMENT     Sequence Label: pAAV-hSyn-GACh2.0
FEATURES             Location/Qualifiers
     source          1..6533
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   47..176
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      627..636
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     633..692
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     protein_bind    696..720
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     1781..1802
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1908..1929)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2169..2188)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    2706..3294
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2759..2779)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3177..3188)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3326..3802
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3525..3544)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3842..3982
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3842..3971
                     /label=AAV2 ITR
     rep_origin      4057..4512
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4144..4163)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4354..4375
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4529..4548)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4648..4670
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        4794..4898
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4899..4967,4968..5759)
                     /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(5117..5136)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5930..6518
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6419..6438
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtcctg caggcagctg
       61 cgcgctcgct cgctcactga ggccgcccgg gcgtcgggcg acctttggtc gcccggcctc
      121 agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg gttcctgcgg
      181 ccgcacgcgt atgaggcggg gtgggggtgc ctacctgacg accgaccccg acccactgga
      241 caagcaccca acccccattc cccaaattgc gcatccccta tcagagaggg ggaggggaaa
      301 caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca ccgcggacag tgccttcgcc
      361 cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa ggcgcgctga cgtcactcgc
      421 cggtcccccg caaactcccc ttcccggcca ccttggtcgc gtccgcgccg ccgccggccc
      481 agccggaccg caccacgcga ggcgcgagat aggggggcac gggcgcgacc atctgcgctg
      541 cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc agcggaggag tcgtgtcgtg
      601 cctgagagcg cagtcgagaa accggtgcca ccatggagac agacacactc ctgctatggg
      661 tactgctgct ctgggttcca ggttccactg gtgacacaag tttgtacaaa aaagttggca
      721 ccatgacctt gcacaataac agtacaacct cgcctttgtt tccaaacatc agctcctcct
      781 ggatacacag cccctccgat gcagggctgc ccccgggaac cgtcactcat ttcggcagct
      841 acaatgtttc tcgagcagct ggcaatttct cctctccaga cggtaccacc gatgaccctc
      901 tgggaggtca taccgtctgg caagtggtct tcatcgcttt cttaacgggc atcctggcct
      961 tggtgaccat catcggcaac atcctggtaa ttgtgtcatt taaggtcaac aagcagctga
     1021 agacggtcaa caactacttc ctcttaagcc tggcctgtgc cgatctgatt atcggggtca
     1081 tttcaatgaa tctgtttacg acctacatca tcatgaatcg atgggcctta gggaacttgg
     1141 cctgtgacct ctggcttgcc attgactacg tagccagcaa tgcctctgtt atgaatcttc
     1201 tggtcatcag ctttgacaga tacttttcca tcacgaggcc gctcacgtac cgagccaaac
     1261 gaacaacaaa gagagccggt gtgatgatcg gtctggcttg ggtcatctcc tttgtccttt
     1321 gggctcctgc catcttgttc tggcaatact ttgttggaaa gagaactgtg cctccgggag
     1381 agtgcttcat tcagttcctc agtgagccca ccattacttt tggcacagcc atcgctgctt
     1441 tttatatgcc tgtcaccatt atgactattt tatactggag ggtctttcag gaggccaaaa
     1501 ggcagctcca gaagattgac aaatctgagg gccgcttcca tgtccagaac cttagccagg
     1561 tggagcaggg aggaaacgtc tatatcaagg ccgacaagca gaagaacggc atcaaggcga
     1621 acttccacat ccgccacaac atcgaggacg gcggcgtgca gctcgcctac cactaccagc
     1681 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcgtgc
     1741 agtccaaact ttcgaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     1801 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagggcggt accggaggga
     1861 gcatggtgag aaagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     1921 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggtgagggc gatgccacct
     1981 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     2041 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     2101 agcagcacga cttcttcaag tccgccatgc ccgaaggcta catccaggag cgcaccatct
     2161 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     2221 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     2281 acaagctgga gtacaacgct cctagcgtcg cagatgggcg gacggggcat ggactccgca
     2341 gatcttccaa gttctgcttg aaggagcaca aagccctcaa gacgctcagt gcgatcttgc
     2401 ttgccttcat catcacttgg accccataca acatcatggt tctggtgaac accttttgtg
     2461 acagctgcat acccaaaacc ttttggaatc tgggctactg gctgtgctac atcaacagca
     2521 ccgtgaaccc cgtgtgctat gctctgtgca acaaaacatt cagaaccact ttcaagatgc
     2581 tgctgctgtg ccagtgtgac aaaaaaaaga ggcgcaagca gcagtaccag cagagacagt
     2641 cggtcatttt tcacaagcgc gcacccgagc aggccttgta ggaattcgat atcaagctta
     2701 tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     2761 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     2821 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     2881 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     2941 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     3001 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     3061 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     3121 tgctcgccta tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
     3181 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
     3241 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat
     3301 accgagcgct gctcgagaga tctacgggtg gcatccctgt gacccctccc cagtgcctct
     3361 cctggccctg gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg
     3421 catcattttg tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg
     3481 gagcaagggg caagttggga agacaacctg tagggcctgc ggggtctatt gggaaccaag
     3541 ctggagtgca gtggcacaat cttggctcac tgcaatctcc gcctcctggg ttcaagcgat
     3601 tctcctgcct cagcctcccg agttgttggg attccaggca tgcatgacca ggctcagcta
     3661 atttttgttt ttttggtaga gacggggttt caccatattg gccaggctgg tctccaactc
     3721 ctaatctcag gtgatctacc caccttggcc tcccaaattg ctgggattac aggcgtgaac
     3781 cactgctccc ttccctgtcc ttctgatttt gtaggtaacc acgtgcggac cgagcggccg
     3841 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     3901 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag
     3961 cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc
     4021 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     4081 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc
     4141 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     4201 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4261 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4321 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4381 cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct
     4441 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     4501 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     4561 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     4621 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     4681 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     4741 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4801 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4861 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4921 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4981 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     5041 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     5101 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     5161 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     5221 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     5281 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     5341 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     5401 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     5461 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     5521 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     5581 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     5641 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5701 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5761 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5821 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5881 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     5941 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     6001 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     6061 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     6121 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     6181 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     6241 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     6301 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     6361 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     6421 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     6481 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcg
//
