LOCUS       sequence-223364-        6227 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For cre-dependent enriched expression of GCaMP6s in axons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6227)
  AUTHORS   Broussard GJ, Liang Y, Fridman M, Unger EK, Meng G, Xiao X, Ji N, 
            Petreanu L, Tian L
  TITLE     In vivo measurement of afferent activity with axon-specific calcium 
            imaging.
  JOURNAL   Nat Neurosci. 2018 Sep;21(9):1272-1280. doi: 
            10.1038/s41593-018-0211-4. Epub 2018 Aug 20.
  PUBMED    30127424
REFERENCE   2  (bases 1 to 6227)
  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/s41593-018-0211-4"; journalName: "Nat Neurosci"; date: 
            "2018-09"; volume: "21"; issue: "9"; pages: "1272-1280"
COMMENT     Sequence Label: pAAV-hSynapsin1-FLEx-axon-GCaMP6s
FEATURES             Location/Qualifiers
     source          1..6227
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     intron          723..855
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     838..857
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     protein_bind    868..901
                     /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    964..997
                     /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(1004..1447,1448..1453,1454..1888,1889..1906,
                     1907..2176,2177..2182,2183..2239,2240..2266,2267..2290,
                     2291..2293,2294..2326,2327..2329,2330..2347,2348..2353))
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="GSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNKT
                     GHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGPV
                     LLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEEL
                     FTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTY
                     GVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIEL
                     KGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSL
                     GQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGNG
                     YISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     1563..1582
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1822..1843
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1949..1970)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      2251..2260
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(2427..2460)
                     /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(2531..2564)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2636..3224
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2689..2709)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3107..3118)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3302..3423
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3313..3332)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3367..3386
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3585..3714
                     /label=AAV2 ITR
     rep_origin      complement(3951..4539)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4031..4050)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4710..5501,5502..5570))
                     /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     5333..5352
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5571..5675)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5701..6156)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5838..5859)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6050..6069
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatcc gagctccagt gtgctggaat tcgccctttt cagtatttaa attgacgacc
      241 gaccccgacc cactggacaa gcacccaacc cccattcccc aaattgcgca tcccctatca
      301 gagaggggga ggggaaacag gatgcggcga ggcgcgtgcg cactgccagc ttcagcaccg
      361 cggacagtgc cttcgccccc gcctggcggc gcgcgccacc gccgcctcag cactgaaggc
      421 gcgctgacgt cactcgccgg tcccccgcaa actccccttc ccggccacct tggtcgcgtc
      481 cgcgccgccg ccggcccagc cggaccgcac cacgcgaggc gcgagatagg ggggcacggg
      541 cgcgaccatc tgcgctgcgg cgccggcgac tcagcgctgc ctcagtctgc ggtgggcagc
      601 ggaggagtcg tgtcgtgcct gagagcgcag ctgtgctcct gggcaccgcg cagtccgccc
      661 ccgcggctcc tggccagacc acccctagga ccccctgccc caagtcgcag ccgttggatc
      721 aggtaagtat caaggttaca agacaggttt aaggagacca atagaaactg ggcttgtcga
      781 gacagagaag actcttgcgt ttctgatagg cacctattgg tcttactgac atccactttg
      841 cctttctctc cacaggtgat gatccccata acttcgtata aagtatccta tacgaagtta
      901 tatcaaaata ggaagaccaa tgcttcacca tcgacccgaa ttgccaagca tcaccatcga
      961 cacataactt cgtataatgt atgctatacg aagttatact agtttacttc gctgtcatca
     1021 tttgtacaaa ctcttcgtag tttacctgac catccccatc gatgtctgct tccctgatca
     1081 tttcatcaac ctcttcatct gttaacttct ctccaaggtt tgtcatcacg tggcgaagct
     1141 ctgctgcact gatgtagcca ttgccatcct tatcaaacac accgaacgct tctctaattt
     1201 cttcttccgt gtccctgtat ttcatttttc ttgccatcat tgtcaggaac tcagggaagt
     1261 cgattgtgcc gtcaccgtcg gcatctactt cattgatcat gtcctgcagc tctgcttctg
     1321 tggggttctg ccccagagac cgcatcaccg tccccagctc cttggttgtt attgtcccat
     1381 ccccgtcctt gtcaaatagg gagaaagcct ctttaaattc tgcgatctgc tcttcagtca
     1441 gttggtccgg caggttgtac tccagcttgt gccccaggat gttgccgtcc tccttgaagt
     1501 cgatgccctt cagctcgatg cggttcacca gggtgtcgcc ctcgaacttc acctcggcgc
     1561 gggtcttgta gttgccgtcg tccttgaaga agatggtgcg ctcctggatg tagccttcgg
     1621 gcatggcgga cttgaagaag tcgtgctgct tcatgtggtc ggggtagcgg ctgaagcact
     1681 gcacgccgta ggtcagggtg gtcacgaggg tgggccaggg cacgggcagc ttgccggtgg
     1741 tgcagatgaa cttcagggtc agcttgccgt aggtggcatc gccctcaccc tcgccggaca
     1801 cgctgaactt gtggccgttt acgtcgccgt ccagctcgac caggatgggc accaccccgg
     1861 tgaacagctc ctcgcccttg ctcaccatgc tccctccggt accgcccttg tacagctcgt
     1921 ccatgccgag agtgatcccg gcggcggtca cgaactccag caggaccatg tgatcgcgct
     1981 tctcgttggg gtctttcgaa agtttggact gcacgctcag gtagtggttg tcgggcagca
     2041 gcacggggcc gtcgccgatg ggggtgttct gctggtagtg gtaggcgagc tgcacgccgc
     2101 cgtcctcgat gttgtggcgg atgtggaagt tcgccttgat gccgttcttc tgcttgtcgg
     2161 ccttgatata gacgttctcg agtgagctca gccgacctat agctctgact gcgtgacctg
     2221 tcttattcca cttacgacgt gatgagtcga ccatggtggc gagatcctta tcgtcatcgt
     2281 cgtacagatc ccgacccatt tgctgtccac cagtcatgct agccatacca tgatgatgat
     2341 gatgatgaga accaatcttt tggtcctcat cattcttttc aacctgtttg gttcttctca
     2401 tacagcacag catggtggcg actagtataa cttcgtatag gatactttat acgaagttat
     2461 cattgggatt cttcctattt tgatccaagc atcaccatcg accctctagt ccagatctca
     2521 ccatcgaccc ataacttcgt atagcataca ttatacgaag ttatgtccct cgaagaggtt
     2581 cgaaacactg aaaagggcga attctgcaga tgatccactc gagaagctta tcgataatca
     2641 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     2701 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     2761 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     2821 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     2881 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     2941 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     3001 cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg
     3061 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     3121 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     3181 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat accgtcgacc
     3241 cgggcggccg ctcgagcatg catctagagt cgacccgggc ggccgcttcg agcagacatg
     3301 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     3361 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     3421 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt
     3481 ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcg ataaggatct tcctagagca
     3541 tggctacgta gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat
     3601 ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt
     3661 cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcat
     3721 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3781 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3841 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3901 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3961 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4021 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4081 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4141 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4201 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4261 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4321 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4381 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4441 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4501 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4561 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4621 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4681 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4741 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4801 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4861 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4921 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4981 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5041 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5101 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5161 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5221 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5281 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5341 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5401 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5461 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5521 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5581 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5641 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5701 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     5761 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     5821 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     5881 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     5941 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     6001 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     6061 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     6121 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
     6181 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccag
//
