LOCUS       sequence-119460-        6730 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ""Fluorescent reporter for calcium imaging"".
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6730)
  AUTHORS   Wertz A, Trenholm S, Yonehara K, Hillier D, Raics Z, Leinweber M, 
            Szalay G, Ghanem A, Keller G, Rozsa B, Conzelmann KK, Roska B
  TITLE     PRESYNAPTIC NETWORKS. Single-cell-initiated monosynaptic tracing 
            reveals layer-specific cortical network modules.
  JOURNAL   Science. 2015 Jul 3;349(6243):70-4. doi: 10.1126/science.aab1687.
  PUBMED    26138975
REFERENCE   2  (bases 1 to 6730)
  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: "10.1126/science"; 
            journalName: "Science"; date: "2015-07-3- 3"; volume: "349"; issue: 
            "6243"; pages: "70-4"
COMMENT     Sequence Label: pAAV-EF1a-GCaMP6s-WPRE-pGHpA
FEATURES             Location/Qualifiers
     source          1..6730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1462..1471
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1468..1476,1477..1494,1495..1497,1498..1530,1531..1533,
                     1534..1557,1558..1584,1585..1641,1642..1647,1648..1917,
                     1918..1935,1936..2370,2371..2376,2377..2820)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      1564..1573
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1854..1875
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1981..2002)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2242..2261)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    2857..3445
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2910..2930)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3328..3339)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3477..3953
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3676..3695)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3993..4133
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3993..4122
                     /label=AAV2 ITR
     rep_origin      4208..4663
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4295..4314)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4505..4526
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4680..4699)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4799..4821
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4859..4877)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4945..5049
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5050..5118,5119..5910)
                     /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(5268..5287)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6081..6669
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6570..6589
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagga
     1441 tcctgtacac cgcgggctag cgccaccatg ggttctcatc atcatcatca tcatggtatg
     1501 gccagcatga ctggtggaca gcaaatgggt cgggatctgt acgacgatga cgataaggat
     1561 ctcgccacca tggtggactc atcacgtcgt aagtggaata agacaggtca cgcagtcaga
     1621 gctataggtc ggctgagctc actcgagaac gtctatatca aggccgacaa gcagaagaac
     1681 ggcatcaagg cgaacttcca catccgccac aacatcgagg acggcggcgt gcagctcgcc
     1741 taccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     1801 tacctgagcg tgcagtccaa actttcgaaa gaccccaacg agaagcgcga tcacatggtc
     1861 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtataagggc
     1921 ggtaccggag ggagcatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
     1981 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggtgag
     2041 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
     2101 gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac
     2161 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacatccag
     2221 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
     2281 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
     2341 aacatcctgg ggcacaagct ggagtacaac ctgccggacc aactgactga agagcagatc
     2401 gcagaattta aagaggcttt ctccctattt gacaaggacg gggatgggac aataacaacc
     2461 aaggagctgg ggacggtgat gcggtctctg gggcagaacc ccacagaagc agagctgcag
     2521 gacatgatca atgaagtaga tgccgacggt gacggcacaa tcgacttccc tgagttcctg
     2581 acaatgatgg caagaaaaat gaaatacagg gacacggaag aagaaattag agaagcgttc
     2641 ggtgtgtttg ataaggatgg caatggctac atcagtgcag cagagcttcg ccacgtgatg
     2701 acaaaccttg gagagaagtt aacagatgaa gaggttgatg aaatgatcag ggaagcagac
     2761 atcgacgggg atggtcaggt aaactacgaa gagtttgtgc aaatgatgac agcgaagtga
     2821 gtcgacagat ctgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt
     2881 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     2941 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     3001 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
     3061 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
     3121 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
     3181 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
     3241 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg
     3301 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
     3361 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
     3421 atctcccttt gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt
     3481 ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc
     3541 ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct
     3601 ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct
     3661 gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca
     3721 ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg
     3781 gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt
     3841 tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc
     3901 ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt
     3961 tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac
     4021 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     4081 gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct
     4141 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa
     4201 ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     4261 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     4321 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     4381 cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt
     4441 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     4501 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggg ctattctttt
     4561 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     4621 aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt
     4681 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     4741 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     4801 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     4861 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     4921 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     4981 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     5041 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5101 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5161 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5221 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     5281 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5341 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5401 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5461 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     5521 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     5581 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     5641 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     5701 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     5761 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     5821 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     5881 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     5941 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6001 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6061 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     6121 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     6181 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     6241 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     6301 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     6361 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     6421 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     6481 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     6541 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     6601 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     6661 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     6721 gctcacatgt
//
