pHybrid-cpGFP-Kir3.2(zebrafish)-mScarlet

LOCUS       pHybrid-cpGFP-Kir3.2(zebrafish)-mScarlet              6302 bp DNA     circular SYN 19-OCT-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pCI-sodium:proton antiporter Bacillus sp. YZJH907-2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6302)
  AUTHORS   admin
  TITLE     Direct Submission
  JOURNAL   Exported Apr 20, 2026 from SnapGene 6.0.2
            https://www.biofeng.com
FEATURES             Location/Qualifiers
     source          1..6302
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        754..872
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     CDS             913..930
                     /codon_start=1
                     /label=6x-HisTag
                     /translation="HHHHHH"
     primer_bind     950..992
                     /label=V-CpGFP-R
     regulatory      961..966
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             970..1692
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="NVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGPV
                     LLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEEL
                     FTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTY
                     GVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIEL
                     KGIDFKEDGNILGHKLEYN"
     primer_bind     complement(1684..1707)
                     /label=CpGFP-R
     primer_bind     1688..1726
                     /label=Kir-zebra-F
     CDS             1693..1707
                     /codon_start=1
                     /label=Link
                     /translation="GGTGG"
     misc_feature    1708..2370
                     /label=Kir3.2-cyto-zebrafish
     primer_bind     complement(2354..2398)
                     /label=Kir-R
     primer_bind     2374..2401
                     /label=rrnB-F
     terminator      2386..2480
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     polyA_signal    2504..2625
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        2640..2884
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2735..2870
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      2885..2890
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2891..3586
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     polyA_signal    3591..3638
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3718..4173
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4505..4609
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4610..5470
                     /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      5641..6229
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tcgtttagtg aaccgtcaga tcactagaag cttcaccaca attcagcaaa ttgtgaacat
      781 catcacgttc atctttccct ggttgccaat ggcccatttt cctgtcagta acgagaaggt
      841 cgcgaattca ggcgcttttt agactggtcg taacccgttt ttggctaaca ggaggaatta
      901 accatgggga gccaccatca ccatcaccat gctagcctcg agaattcacg cgtggtacca
      961 gccaccatga acgtctatat caaggccgac aagcagaaga acggcatcaa ggcgaacttc
     1021 aagatccgcc acaacatcga ggacggcggc gtgcagctcg cctaccacta ccagcagaac
     1081 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cgtgcagtcc
     1141 aaactttcga aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     1201 gccgccggga tcactctcgg catggacgag ctgtacaagg gcggtaccgg agggagcatg
     1261 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     1321 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggtg agggcgatgc cacctacggc
     1381 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1441 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     1501 cacgacttct tcaagtccgc catgcccgaa ggctacatcc aggagcgcac catcttcttc
     1561 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     1621 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     1681 ctggagtaca acggcggcac cggcggcaag aagagagccg agaccctggt gttcagcacc
     1741 aacgccgtga tcagcatgag agacggaaga ctgtgcctga tgttccgggt gggagacctg
     1801 agaaacagcc acatcgtgga ggccagcatc agagccaaac tgatcaagag taaacagacc
     1861 aaagagggag aattcatccc cctgaaccag accgacatca acgtgggata cgacaccgga
     1921 gacgacagac tgttcctggt gagccccctg atcatctgcc acgaaatcaa ccagcacagc
     1981 ccattctggg agatctcccg cgaacacatg gccaaagaag agctggagat cgtggtgatt
     2041 ctggagggca tggtggaggc caccggcatg acctgtcagg ccaggagcag ctatgtggcc
     2101 agcgagatca agtggggcta cagattcacc cccgtgctga ccctggagga tgggttttat
     2161 gaggtggact ataactcctt ccacgagatc tacgagacca acacacccag ctgctccgcc
     2221 agagagctgg ccgaaatgac cgccagagcc cggctgcccc tgacctggtc agtggctagc
     2281 aagctgagcc agctggagcc ccaggaaccc ccagaagaac tgaccgaaag aaacggagac
     2341 gtggccaaca tggagagcga aagcaaggtg tagtctagag tcgacccagg catcaaataa
     2401 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     2461 ctctcctgag taggacaaat gcggccgctt cgagcagaca tgataagata cattgatgag
     2521 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     2581 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgg
     2641 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2701 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     2761 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     2821 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     2881 caaagccacc atggtgagca agggcgaggc agtgatcaag gagttcatgc ggttcaaggt
     2941 gcacatggag ggctccatga acggccacga gttcgagatc gagggcgagg gcgagggccg
     3001 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt
     3061 ctcctgggac atcctgtccc ctcagttcat gtacggctcc agggccttca ccaagcaccc
     3121 cgccgacatc cccgactact ataagcagtc cttccccgag ggcttcaagt gggagcgcgt
     3181 gatgaacttc gaggacggcg gcgccgtgac cgtgacccag gacacctccc tggaggacgg
     3241 caccctgatc tacaaggtga agctccgcgg caccaacttc cctcctgacg gccccgtaat
     3301 gcagaagaag acaatgggct gggaagcgtc caccgagcgg ttgtaccccg aggacggcgt
     3361 gctgaagggc gacattaaga tggccctgcg cctgaaggac ggcggccgct acctggcgga
     3421 cttcaagacc acctacaagg ccaagaagcc cgtgcagatg cccggcgcct acaacgtcga
     3481 ccgcaagttg gacatcacct cccacaacga ggactacacc gtggtggaac agtacgaacg
     3541 ctccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtgac cggcaataaa
     3601 aagacagaat aaaacgcacg gtgttgggtc gtttgttcgg atccgggctg gcgtaatagc
     3661 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggacg
     3721 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     3781 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     3841 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     3901 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     3961 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4021 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     4081 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4141 cgaattttaa caaaatatta acgcttacaa tttcctgatg cggtattttc tccttacgca
     4201 tctgtgcggt atttcacacc gcatatggtg cactctcagt acaatctgct ctgatgccgc
     4261 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     4321 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     4381 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
     4441 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     4501 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     4561 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     4621 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     4681 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     4741 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     4801 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     4861 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     4921 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     4981 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5041 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5101 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5161 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5221 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     5281 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     5341 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     5401 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     5461 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     5521 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     5581 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     5641 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     5701 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     5761 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     5821 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     5881 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     5941 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6001 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6061 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6121 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6181 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6241 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgg ctcgacagat
     6301 ct
//