LOCUS       sequence-193740-        5929 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV expression of Twitch2B driven from hSyn1 promoter. Twitch 2B is 
            a fluorescent reporter for calcium signaling .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5929)
  AUTHORS   Thestrup T, Litzlbauer J, Bartholomaus I, Mues M, Russo L, Dana H, 
            Kovalchuk Y, Liang Y, Kalamakis G, Laukat Y, Becker S, Witte G, 
            Geiger A, Allen T, Rome LC, Chen TW, Kim DS, Garaschuk O, Griesinger
            C, Griesbeck O
  TITLE     Optimized ratiometric calcium sensors for functional in vivo imaging
            of neurons and T lymphocytes.
  JOURNAL   Nat Methods. 2014 Jan 5. doi: 10.1038/nmeth.2773.
  PUBMED    24390440
REFERENCE   2  (bases 1 to 5929)
  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: "Nat 
            Methods. 2014 Jan 5. doi: 10.1038/nmeth.2773."
COMMENT     Sequence Label: pAAV.hSyn1.Twitch2B.WPRE.SV40
FEATURES             Location/Qualifiers
     source          1..5929
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        157..604
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      613..622
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(664..685)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(925..944)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    2312..2900
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2365..2385)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2783..2794)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2903..2919)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2904..2920)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2949..3070
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2960..2979)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3014..3033
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3232..3361
                     /label=AAV2 ITR
     rep_origin      complement(3598..4186)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3678..3697)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4357..5148,5149..5217))
                     /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     4980..4999
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5218..5322)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5348..5803)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5485..5506)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5697..5716
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   join(5875..5929,1..75)
                     /label=AAV2 ITR
ORIGIN
        1 cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg gccaactcca
       61 tcactagggg ttccttgtag ttaatgatta acccgccatg ctacttatct acgtagccat
      121 gctctaggaa gatctctgca gagggccctg cgtatgagtg caagtgggtt ttaggaccag
      181 gatgaggcgg ggtgggggtg cctacctgac gaccgacccc gacccactgg acaagcaccc
      241 aacccccatt ccccaaattg cgcatcccct atcagagagg gggaggggaa acaggatgcg
      301 gcgaggcgcg tgcgcactgc cagcttcagc accgcggaca gtgccttcgc ccccgcctgg
      361 cggcgcgcgc caccgccgcc tcagcactga aggcgcgctg acgtcactcg ccggtccccc
      421 gcaaactccc cttcccggcc accttggtcg cgtccgcgcc gccgccggcc cagccggacc
      481 gcaccacgcg aggcgcgaga taggggggca cgggcgcgac catctgcgct gcggcgccgg
      541 cgactcagcg ctgcctcagt ctgcggtggg cagcggagga gtcgtgtcgt gcctgagagc
      601 gcagtcgaat tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
      661 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
      721 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
      781 cccgtgccct ggcccaccct cgtgaccacc ctgagctggg gcgtgcagtg cttcgcccgc
      841 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
      901 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
      961 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
     1021 ggcaacatcc tggggcacaa gctggagtac aacgccatcc acggcaacgt ctatatcacc
     1081 gccgacaagc agaagaacgg catcaaggcc aacttcggcc tcaactgcaa catcgaggac
     1141 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
     1201 ctgctgcccg acaaccacta cctgagcacc cagtccaagc tgagcaaaga ccccaacgag
     1261 aagcgcgatc acatggtcct gctggagttc gtgaccgccg cccgcatgca agttgccgac
     1321 gccagcgaag aggaactgag cgagtgcttc agaatcttcg acttcgacgg caacggcttc
     1381 atcgacagag aggagtttgg cgacatcatc agactgaccg gcgagcagct gaccgacgag
     1441 gacgtggacg agatcttcgg cgactccgac accgacaaga acggcagaat cgatttcgac
     1501 gagttcctga agatggtgga aaacgtgcag cccatctacc ccgagctcat gggtggggtc
     1561 cagcttgcag atcactacca gcagaatact ccgatcgggg atgggccggt tctcctgcct
     1621 gacaaccact acctgtcata ccagtccaag ctttccaaag atcctaatga aaagcgggat
     1681 cacatggtcc tgctggaatt tgtcaccgcc gccggaatca ctctcggaat ggacgagctg
     1741 tacaaaggtg ggaccggggg atctatggtg agcaagggcg aagaattgtt cactggtgta
     1801 gtccctattt tggtggaact ggatggggat gtcaacggtc ataaattcag cgtgcgaggg
     1861 gagggtgagg gggacgcaac taatggcaaa ttgacactca agctcatctg cacaacgggg
     1921 aaactgcccg tgccatggcc cacactcgtt actactctag gatatggcct tatgtgtttc
     1981 gcccgctacc cagaccatat gaaacaacac gatttcttta aatcagccat gccagagggg
     2041 tacgtccagg agagaactat cttcttcaag gacgatggta attacaagac tagggccgag
     2101 gtgaaattcg aaggggatac acttgtcaac cgaatcgagc tgaagggaat cgactttaag
     2161 gaggacggaa acatactcgg gcataagttg gaatacaact acaacagtca caatgtttac
     2221 atcacagccg ataagcaaaa gaacggcatt aaggctaatt ttaagatccg tcacaacatt
     2281 gaggattgat gatcaactag agcttatcga taatcaacct ctggattaca aaatttgtga
     2341 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     2401 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     2461 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     2521 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     2581 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     2641 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     2701 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     2761 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     2821 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     2881 cctttgggcc gcctccccgc atcgataccg tcgacctcga cccgggcggc cgcttcgagc
     2941 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     3001 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     3061 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggagatgtg
     3121 ggaggttttt taaagcaagt aaaacctcta caaatgtggt aaaatcgata aggatcttcc
     3181 tagagcatgg ctacgtagat aagtagcatg gcgggttaat cattaactac aaggaacccc
     3241 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     3301 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     3361 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     3421 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     3481 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     3541 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3601 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3661 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3721 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3781 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3841 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3901 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3961 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4021 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4081 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4141 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4201 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4261 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4321 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     4381 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     4441 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     4501 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     4561 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     4621 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     4681 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     4741 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     4801 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     4861 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     4921 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     4981 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5041 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5101 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5161 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5221 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5281 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     5341 gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     5401 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     5461 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     5521 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     5581 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     5641 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     5701 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     5761 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg
     5821 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctgcgc
     5881 gctcgctcgc tcactgaggc cgcccgggca aagcccgggc gtcgggcga
//
