LOCUS       sequence-82296-e        5929 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for calcium signaling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5929)
  TITLE     pZac2.1gfaABC1Dlck-GCaMP6f
  JOURNAL   Unpublished
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: 
            "Unpublished"
COMMENT     Sequence Label: pZac2.1 gfaABC1D-lck-GCaMP6f
FEATURES             Location/Qualifiers
     source          1..5929
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   259..388
                     /label=AAV2 ITR
     intron          1239..1371
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1354..1373
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1416..1434
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1556..1565
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1562..1570,1571..1588,1589..1591,1592..1624,1625..1627,
                     1628..1651,1652..1678,1679..1735,1736..1741,1742..2011,
                     2012..2029,2030..2464,2465..2470,2471..2914)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      1658..1667
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1948..1969
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2075..2096)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2336..2355)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    2954..3075
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2965..2984)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3019..3038
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3237..3366
                     /label=AAV2 ITR
     primer_bind     complement(3385..3402)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3385..3401)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3394..3416)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3543..3998
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3630..3649)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3840..3861
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4024..4128
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4129..4197,4198..4989)
                     /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"
     rep_origin      5160..5748
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5649..5668
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccagattta
      241 attaaggcct taattaggct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc
      301 gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga
      361 gtggccaact ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta
      421 tctacgtagc catgctctag gaagatctaa catatcctgg tgtggagtag gggacgctgc
      481 tctgacagag gctcgggggc ctgagctggc tctgtgagct ggggaggagg cagacagcca
      541 ggccttgtct gcaagcagac ctggcagcat tgggctggcc gccccccagg gcctcctctt
      601 catgcccagt gaatgactca ccttggcaca gacacaatgt tcggggtggg cacagtgcct
      661 gcttcccgcc gcaccccagc ccccctcaaa tgccttccga gaagcccatt gagcaggggg
      721 cttgcattgc accccagcct gacagcctgg catcttggga taaaagcagc acagccccct
      781 aggggctgcc cttgctgtgt ggcgccaccg gcggtggaga acaaggctct attcagcctg
      841 tgcccaggaa aggggatcag gggatgccca ggcatggaca gtgggtggca gggggggaga
      901 ggagggctgt ctgcttccca gaagtccaag gacacaaatg ggtgagggga gagctctccc
      961 catagctggg ctgcggccca accccacccc ctcaggctat gccagggggt gttgccaggg
     1021 gcacccgggc atcgccagtc tagcccactc cttcataaag ccctcgcatc ccaggagcga
     1081 gcagagccag agcaggttgg agaggagacg catcacctcc gctgctcgca agctttattg
     1141 cggtagttta tcacagttaa attgctaacg cagtcagtgc ttctgacaca acagtctcga
     1201 acttaagctg cagaagttgg tcgtgaggca ctgggcaggt aagtatcaag gttacaagac
     1261 aggtttaagg agaccaatag aaactgggct tgtcgagaca gagaagactc ttgcgtttct
     1321 gataggcacc tattggtctt actgacatcc actttgcctt tctctccaca ggtgtccact
     1381 cccagttcaa ttacagctct taaggctaga gtacttaata cgactcacta taggctagcc
     1441 tcgagaattc accatgggct gtggctgcag ctcaaaccct gaagatgact ggatggagaa
     1501 cattgacgtg tgtgagaact gccattatcc cccgccgaag cttcgaatgg atccagccac
     1561 catgggttct catcatcatc atcatcatgg tatggctagc atgactggtg gacagcaaat
     1621 gggtcgggat ctgtacgacg atgacgataa ggatctcgcc accatggtcg actcatcccg
     1681 tcgtaagtgg aataagacag gtcacgcagt cagagctata ggtcggctga gctcactcga
     1741 gaacgtctat atcaaggccg acaagcagaa gaacggcatc aaggcgaact tcaagatccg
     1801 ccacaacatc gaggacggcg gcgtgcagct cgcctaccac taccagcaga acacccccat
     1861 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcgtgcagt ccaaactttc
     1921 gaaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     1981 gatcactctc ggcatggacg agctgtacaa gggcggtacc ggagggagca tggtgagcaa
     2041 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     2101 cggccacaag ttcagcgtgt ccggcgaggg tgagggcgat gccacctacg gcaagctgac
     2161 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     2221 cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt
     2281 cttcaagtcc gccatgcccg aaggctacat ccaggagcgc accatcttct tcaaggacga
     2341 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     2401 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     2461 caacctgccg gaccaactga ctgaagagca gatcgcagaa tttaaagagg aattctccct
     2521 atttgacaag gacggggatg ggacaataac aaccaaggag ctggggacgg tgatgcggtc
     2581 tctggggcag aaccccacag aagcagagct gcaggacatg atcaatgaag tagatgccga
     2641 cggtgacggc acaatcgact tccctgagtt cctgacaatg atggcaagaa aaatgaaata
     2701 cagggacacg gaagaagaaa ttagagaagc gttcggtgtg tttgataagg atggcaatgg
     2761 ctacatcagt gcagcagagc ttcgccacgt gatgacaaac cttggagaga agttaacaga
     2821 tgaagaggtt gatgaaatga tcagggaagc agacatcgat ggggatggtc aggtaaacta
     2881 cgaagagttt gtacaaatga tgacagcgaa gtgatctaga gtcgacccgg gcggccgctt
     2941 cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     3001 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     3061 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     3121 atgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat cgataaggat
     3181 cttcctagag catggctacg tagataagta gcatggcggg ttaatcatta actacaagga
     3241 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
     3301 gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc
     3361 gcgcagcctt aattaaccta attcactggc cgtcgtttta caacgtcgtg actgggaaaa
     3421 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     3481 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     3541 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     3601 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     3661 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     3721 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     3781 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     3841 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     3901 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     3961 taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat
     4021 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     4081 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     4141 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     4201 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     4261 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     4321 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     4381 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     4441 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     4501 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     4561 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     4621 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     4681 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     4741 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     4801 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     4861 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     4921 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     4981 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     5041 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     5101 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     5161 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     5221 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     5281 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     5341 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     5401 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     5461 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     5521 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     5581 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     5641 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     5701 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     5761 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     5821 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     5881 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaag
//
