LOCUS       sequence-421945-        7630 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ubiquitously expresses jGCaMP7f in zebrafish via tol2 transgenesis 
            or in vitro transcription of mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7630)
  AUTHORS   Jia BZ, Qi Y, Wong-Campos JD, Megason SG, Cohen AE
  TITLE     A bioelectrical phase transition patterns the first vertebrate 
            heartbeats.
  JOURNAL   Nature. 2023 Oct;622(7981):149-155. doi: 10.1038/s41586-023-06561-z.
            Epub 2023 Sep 27.
  PUBMED    37758945
REFERENCE   2  (bases 1 to 7630)
  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/s41586-023-06561-z"; journalName: "Nature"; date: 
            "2023-10"; volume: "622"; issue: "7981"; pages: "149-155"
COMMENT     Sequence Label: pBJ053_MTB_jGCaMP7f
FEATURES             Location/Qualifiers
     source          1..7630
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    join(13..24,25..212)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     promoter        2928..2946
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2928..2945
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2954..2973
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2954..2972
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      3004..3013
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3010..3018,3019..3036,3037..3039,3040..3072,3073..3075,
                     3076..3099,3100..3126,3127..3183,3184..3189,3190..3459,
                     3460..3477,3478..3912,3913..3918,3919..4362)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKELFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYTDSEEEIREAFRVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      3106..3115
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     3396..3417
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3523..3544)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3784..3803)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    4388..4522
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4412..4431)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4466..4485
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(4614..4751,4752..4763)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(4822..4839)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4822..4838)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4831..4853)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4997..5016)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5207..5228
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5391..5495
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5496..5564,5565..6356)
                     /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(5714..5733)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6527..7115
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7016..7035
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7269..7286
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7403..7424
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7439..7444,7445..7462,7463..7469)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7477..7493
                     /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     7482..7504
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7501..7517
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7501..7517
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7536..7556
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7538..7556
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     7593..7609
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7593..7609
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
ORIGIN
        1 ggggtcaaga accagaggtg taaagtactt gagtaatttt acttgattac tgtacttaag
       61 tattattttt ggggattttt actttacttg agtacaatta aaaatcaata cttttacttt
      121 tacttaatta cattttttta gaaaaaaaag tactttttac tccttacaat tttatttaca
      181 gtcaaaaagt acttattttt tggagatcac ttgggcccgg ctcgagggcc tagtcattac
      241 ctatactcaa cgtttactct gagaaacttc catattttag aggtaaaatg ctcagaattg
      301 taaaaagtct tgtcaaaata cttggaattt gccaaaaaca aaataaaagt cacatcacct
      361 cattgtcctt ggtatcagca agcactttta acacaatcag ctgtttttgt ttataactcc
      421 cactgaaggc catgtgcatt actcccaagt tacacagtaa aaagtcacat gtagtttatc
      481 atgatcactg ctgctcaagc acagctttat ctgttattca gtgtggcaat atctgaaatg
      541 tgtgcaaaat attatttatt gtagctgatc accaacactt tcagtgtttt tttttttcta
      601 cttttaatga gaattatgtt cttaggattt atccttataa acctctttaa aaagaaatgt
      661 tatgtctgca tagacacgac atgaaattat gaacagaaaa tttaactatt taagatcaag
      721 acttgacata taacatttct aatgaacatt aaacctgagg taggcccact tgcaaaaatg
      781 taactgctat cactcacttt atacatcatt gtgtaacttg tacacacaaa aatagtgaag
      841 caatgtggca tggaatgcag gtcccattca ttcaaagtta ttaaagcact ttcttaaagt
      901 gcattactac tgagctaaga ccagctagtt gctcatacaa catactacaa gtagtgtcat
      961 cctaatttgg ataaaaaaaa tgtccatttt atacatggaa aagtactcat tagatccccc
     1021 attgtttgta ttacggtatt tcgtgaacac aagaggtaaa tgacctacag agctgctgtt
     1081 gtgttagatt gaaaacacaa cacaggatca tggaggcatc catcattatg acgcattctg
     1141 ccacttgtaa ctctgcacag actttttgaa aagtttaatt gagtcttggc gttgtcgcag
     1201 aaaaatacga aaaattctgt gttaaaattt gcaaataaaa attccagttt tataggaaat
     1261 cgccatggcc ttgttcttta accagttaag cttccccttc tttcactctc aagttgcaag
     1321 aagcaagtgt agcaatgtgc acgcgacagc cgggtgtgtg acgctggacc aatcagagca
     1381 cagagctccg aaagtttacc ttttatggct agagccgggt atgtgccgtc atataaaaga
     1441 gcgcgcccag cttttcagcc tcactttgag ctcctccaca cgcagctagt gcggaatatc
     1501 atctgcttgt aacccattct cttaagtcga caacccccca aacccaaggt gagttggttt
     1561 tacagttttt aagcgtttaa ttatagtttc tatctattaa tgttaagtaa aatgtgtaaa
     1621 tggataaata tgatttgatg aagtctttag acagttcata acggtctctt tgtagttatc
     1681 tactaggttt gatttataag tggtggttct tttgataatc tatgttatat ttaataaata
     1741 ttggctgtat cttaactttt ttgtagcgtt agtggatgtg gcaggtgaga atgtcggtaa
     1801 tttaacgtga ccagtcgtag gcacgacatt gaacattgaa taggccggtg tgaaatgggt
     1861 gttaagtctt aatataaagt tgtgttctgg ctaatctctc tttaacagcc ataaatgtga
     1921 tttttaacta cagttttgag ggtgctggtg agcatgttgc acacttgatg gatagccggc
     1981 atgggaagtt ctttgtgcag gcagtgctgc agcagggtgt gacctacttt agctagccgg
     2041 ctaaccagct ctcatctgat gtaacgtaaa ccccatcaat ctaacatcgg ctcattcgtg
     2101 ctttataaga tcgaagatct tgttattagt aacgggtatt ggcatttagt tttaacttgg
     2161 ttaattttct ttttgttatg tgttctattt aaatgtagcc tgcagccccc tctgtcttta
     2221 atgaagctga agattaacgt taacgttagt gaagtgttgt atccgctatg aaagatttga
     2281 attttagagg tgtggtccac tttaataacg gctgcggagt tctctggctg cacttcagcc
     2341 gctgtgactg tcgaagcaga gcccatcagt ttttcaggat ctcggtgatt tttgaggcta
     2401 gactatgaac tgaaccgact gagagtgacc gtccagaaat gctttagtgt gaagtctccg
     2461 ctcacgagga gtccgtggac tgactgcaga tctgtgacgc agtaacaccc ggcggcagac
     2521 acccgttgaa atgcggttgt gtaattgatc gcaggcgagg gtggaagagg atgtgaaact
     2581 tcatttgtgt aaaatttagg gagtggtcct ggtgtgatga atgtcgaaat ctgttccttt
     2641 ttactgagcc ctacgactct ggctgagtgc cacaccgccg gcagccgcaa agcggctcaa
     2701 accattgcct tttatggtaa taatgagaga atgcagaggg acttcctttg tctggcatat
     2761 ctgaggcgcg cgctgtcact agcgcccacc agcggtcaga ctgtagaatg cagagcaaaa
     2821 caggaagttg actccagatg gtcacatgct tgctgaaacg cctctccctg gcagcagtgc
     2881 aattctaaca gtttcccttt cttttacagt tcagccatcc taggcctatt taggtgacac
     2941 tatagaagag tactaatacg actcactata gggagaagct acttgttctt tttgcaggat
     3001 cccgccacca tgggttctca tcatcatcat catcatggta tggctagcat gactggtgga
     3061 cagcaaatgg gtcgggatct gtacgacgat gacgataagg atctcgccac catggtcgac
     3121 tcatcacgtc gtaagtggaa taagacaggt cacgcagtca gagctatagg tcggctgagc
     3181 tcactcgaga acgtctatat caaggccgac aagcagaaga acggcatcaa ggcgaacttc
     3241 aagatccgcc acaacatcga ggacggcggc gtgcagctcg cctaccacta ccagcagaac
     3301 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cgtgcagtcc
     3361 aaactttcga aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     3421 gccgccggga tcactctcgg catggacgag ctgtacaagg gcggtaccgg agggagcatg
     3481 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     3541 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggtg agggcgatgc cacctacggc
     3601 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     3661 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     3721 cacgacttct tcaagtccgc catgcccgaa ggctacatcc aggagcgcac catcttcttc
     3781 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     3841 aaccgcatcg agcttaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     3901 ctggagtaca acctgccgga ccaactgact gaagagcaga tcgcagaatt taaagagctg
     3961 ttctccctat ttgacaagga cggggatggg acaataacaa ccaaggagct ggggacggtg
     4021 atgcggtctc tggggcagaa ccccacagaa gcagagctgc aggacatgat caatgaagta
     4081 gatgccgacg gtgacggcac aatcgacttc cctgagttcc tgacaatgat ggcaagaaaa
     4141 atgaaataca cagacagtga agaagaaatt agagaagcgt tccgtgtgtt tgataaggat
     4201 ggcaatggct acatcagtgc agcagagctt cgccacgtga tgacaaacct tggagagaag
     4261 ttaacagatg aagaggttga tgaaatgatc agggaagcag acatcgatgg ggatggtcag
     4321 gtaaactacg aagagtttgt acaaatgatg acagcgaagt gagaattcat acgtatccgg
     4381 aaccggtgat ccagacatga taagatacat tgatgagttt ggacaaacca caactagaat
     4441 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     4501 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     4561 gggggaggtg tgggaggttt tttgatatcc cgggtttaaa cgcatgcaga tctaatactc
     4621 aagtacaatt ttaatggagt acttttttac ttttactcaa gtaagattct agccagatac
     4681 ttttactttt aattgagtaa aattttccct aagtacttgt actttcactt gagtaaaatt
     4741 tttgagtact ttttacacct ctgtcaagaa ccatatgccg gtacccaatt cgccctatag
     4801 gacgtcgtat tacgcgcgct cactggccgt cgttttacaa cgtcgtgact gggaaaaccc
     4861 tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag
     4921 cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggga
     4981 cgcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     5041 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     5101 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     5161 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     5221 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     5281 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     5341 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
     5401 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     5461 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     5521 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     5581 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     5641 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     5701 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     5761 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     5821 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     5881 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     5941 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6001 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     6061 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     6121 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     6181 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     6241 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6301 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     6361 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     6421 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     6481 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     6541 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     6601 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     6661 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     6721 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     6781 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6841 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     6901 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     6961 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     7021 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     7081 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     7141 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     7201 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     7261 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     7321 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     7381 cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt
     7441 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
     7501 caggaaacag ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac
     7561 aaaagctgga gctccaccgc ggtggcggcc gctctagaac tagtggatct gctgggcttg
     7621 ctgaaggtag
//
