LOCUS       sequence-339084-        7054 bp    DNA     circular SYN 14-APR-2025
DEFINITION  TnC-based calcium sensor with green-red FRET pair ratiometric 
            readout.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7054)
  AUTHORS   Zhang D, Redington E, Gong Y
  TITLE     Rational engineering of ratiometric calcium sensors with bright 
            green and red fluorescent proteins.
  JOURNAL   Commun Biol. 2021 Jul 29;4(1):924. doi: 10.1038/s42003-021-02452-z.
  PUBMED    34326458
REFERENCE   2  (bases 1 to 7054)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Commun 
            Biol."; date: "2021-07-29"; volume: "4(1):924. doi"; pages: " 
            10.1038/s42003-021-02452-z"
COMMENT     Sequence Label: AAV-Camk2-NES-Twitch-GR
FEATURES             Location/Qualifiers
     source          1..7054
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   79..219
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   90..219
                     /label=AAV2 ITR
     polyA_signal    259..735
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     517..536
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    767..1355
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             873..884
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1282..1302
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     primer_bind     complement(2589..2608)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2858..2876
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     promoter        complement(3099..4387)
                     /label=CaMKII promoter
                     /note="promoter for the mouse calcium/calmodulin-dependent 
                     kinase II alpha subunit"
     repeat_region   4406..4535
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(4597..5185)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4677..4696)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5356..6147,6148..6216))
                     /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     5979..5998
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6217..6321)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6389..6407
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6445..6467)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6567..6586
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(join(6603..7054,1..4))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6740..6761)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6952..6971
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gcgtactatg gttgctttga cgtatgcggt gtgaaatacc gcacagatgc gtaaggagaa
       61 aataccgcat caggcgcccc tgcaggcagc tgcgcgctcg ctcgctcact gaggccgccc
      121 gggcaaagcc cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc
      181 gcagagaggg agtggccaac tccatcacta ggggttcctg cggccgctcg gtccgcacgt
      241 ggttacctac aaaatcagaa ggacagggaa gggagcagtg gttcacgcct gtaatcccag
      301 caatttggga ggccaaggtg ggtagatcac ctgagattag gagttggaga ccagcctggc
      361 caatatggtg aaaccccgtc tctaccaaaa aaacaaaaat tagctgagcc tggtcatgca
      421 tgcctggaat cccaacaact cgggaggctg aggcaggaga atcgcttgaa cccaggaggc
      481 ggagattgca gtgagccaag attgtgccac tgcactccag cttggttccc aatagacccc
      541 gcaggcccta caggttgtct tcccaacttg ccccttgctc cataccaccc ccctccaccc
      601 cataatatta tagaaggaca cctagtcaga caaaatgatg caacttaatt ttattaggac
      661 aaggctggtg ggcactggag tggcaacttc cagggccagg agaggcactg gggaggggtc
      721 acagggatgc cacccgtaga tctctcgagc agcgctcggt atcgatgcgg ggaggcggcc
      781 caaagggaga tccgactcgt ctgagggcga aggcgaagac gcggaagagg ccgcagagcc
      841 ggcagcaggc cgcgggaagg aaggtccgct ggattgaggg ccgaagggac gtagcagaag
      901 gacgtcccgc gcagaatcca ggtggcaaca taggcgagca gccaaggaaa ggacgatgat
      961 ttccccgaca acaccacgga attgtcagtg cccaacagcc gagcccctgt ccagcagcgg
     1021 gcaaggcagg cggcgatgag ttccgccgtg gcaataggga gggggaaagc gaaagttccg
     1081 gaaaggagct gacaggtggt ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca
     1141 gtgcacacca cgccacgttg cctgacaacg ggccacaact cctcataaag agacagcaac
     1201 caggatttat acaaggagga gaaaatgaaa gccatacggg aagcaatagc atgatacaaa
     1261 ggcattaaag cagcgtatcc acatagcgta aaaggagcaa catagttaag aataccagtc
     1321 aatctttcac aaattttgta atccagaggt tgattatcga taagcttgat atcgaattca
     1381 ttaaccatct tcgacattat gtcggatctt aaagttggcc tttatcccat ttttctgctt
     1441 atcagcggta atatacacat tatggctgtt aaagttgtac tccaacttgt gccccagtat
     1501 gttaccgtcc tccttaaaat caataccttt cagttcaatt cgattaacca gagtgtcccc
     1561 ttcaaattta acctcggctc gggttttata tgtaccatca tccttaaagc tgatggtcct
     1621 ttcttgaaca taaccttcag gcattgcaga cttgaaaaag tcatgctgtt tcatgtggtc
     1681 gggatatcga gaaaagcatt gcacgccgta ggtgagggtc gttacgagtg taggccatgg
     1741 gacgggcagt ttgcccgtcg tacagatgaa tttcagagtc agcttaccat tcgtggcgtc
     1801 cccttcgcct tcgcccctga cggagaattt atgtccattt acatctccat caagctctac
     1861 cagaatgggc actactccag tgaagagttc ttcaccttta ctcaccatgc tgccgccggt
     1921 acctcccttg tacagctcat ccataccgag agtgatgcct gcggcagtca caaattccag
     1981 aaggaccatg tgatctctct tttcgtttgg gtccttggaa agctttgatt gcgtggagag
     2041 ataatgattg tcaggcaaga gtacaggtcc atctccgatg ggcgtatttt gttggtaatg
     2101 atcagccagt tgaacactgg ccagaggctg gacattctcc accatcttca ggaactcatc
     2161 aaagtcaatc cttccatttt tgtctgtgtc tgaatctcca aatatctcat cgacatcttc
     2221 atctgtgagc tgttctccag taaggcggat gatgtctcca aattcctccc tgtcgatgaa
     2281 accatttcca tcgaagtcaa aaatgcgaaa gcactctgat agctcttctt cgctagcatc
     2341 ggagtggcgg ccctcggagc gttcgtactg ttccaccacg gtgtagtcct cgttgtggga
     2401 ggtgatgtcc aacttgcggt cgacgttgta ggcgccgggc atctgcacgg gcttcttggc
     2461 cttgtaggtg gtcttgaagt ccgccaggta gcggccgccg tccttcaggc gcagggccat
     2521 cttaatgtcg cccttcagca cgccgtcctc ggggtacaac cgctcggtgg acgcttccca
     2581 gcccattgtc ttcttctgca ttacggggcc gtcaggaggg aagttggtgc cgcggagctt
     2641 caccttgtag atcagggtgc cgtcctccag ggaggtgtcc tgggtcacgg tcacggcgcc
     2701 gccgtcctcg aagttcatca cgcgctccca cttgaagccc tcggggaagg actgcttata
     2761 gtagtcgggg atgtcggcgg ggtgcttggt gaaggccctg gagccgtaca tgaactgagg
     2821 ggacaggatg tcccaggaga agggcagggg gccacccttg gtcaccttca gcttggcggt
     2881 ctgggtgccc tcgtaggggc ggccctcgcc ctcgccctcg atctcgaact cgtggccgtt
     2941 catggagccc tccatgtgca ccttgaaccg catgaactcc ttgatcactg cctcgccctt
     3001 gctcacagaa cctcctccag tcttgttaat atcaagtcca gccaacttaa gagcaagctc
     3061 gttctgcagc atggtggcgg atccggtacc gctctagagc tgcccccaga actaggggcc
     3121 actcgcctgc ccgtgctcct gagtgcaaac ggagaaccgg cttgactgac gagcttgggg
     3181 cttctgagca gggcactgtg gctgctcaca gcctcgcgag ccttcgtcag catccaggtc
     3241 cccatggcaa ccacctccga aacgccctgt tcccgttgcc ccggtaactg cctccccaac
     3301 acctgcctgc ctttcacttt aaaacctgct cctctttgcc ctggccttat atacacaata
     3361 ctgcaggact gtgtgggccc aggagcaagt ggaccctgtt cccccagggt gatgtactga
     3421 gggggttgga gaaggggatg cgaacaaact tagtccacaa gtgatcatcg ctcttcctac
     3481 ctcccccacc tccaactata gggccctttt aacagacggg atgggattaa gtggggcaaa
     3541 gaggcagttg ctatggtaac ggctagtggg tgccacattc tgggatggtc cttgagaaag
     3601 acaggatgta gttaccctgg caacttcatc tcctctgaag agacaggtat tgccccgttg
     3661 ccctggcaac agctgggttc taaggtttac aggggaagct aattcatctc tccttccctt
     3721 cccccttgta gtctctgtaa tggtttcttt gcttcctctg ggaattggtc tggttgactt
     3781 ggcctctgtc tctgtccttg tttctttcca gctctccctg ctttttagcc attgtgcata
     3841 tcacaccgcc accttagcgt gagaagaagt accaaacaga cccagaatgc tgacggaatg
     3901 actctgtctg ctttctcacc cctatctgtc accactgagc agcagggccc ctcctgaggc
     3961 ctctgccaca gctctcccca cccatcctcc cagagggacc atgaagattt ggtctcctgg
     4021 ggctcctgtc ccagtttctg cttgtgcttc ccgtcaaggt aggactggag agaggcaact
     4081 ataatgaggt gaacgtggct ggactcaccc agggctgggc gggtggtgta gatgtaccct
     4141 gccctggact cttctttcct aaactcattg gtcatctgcc ctaatgagtt cactccttct
     4201 gtatatcgta cttgcatgcc cccaaggcat ggaatttaag attagtaatc tgagttcaag
     4261 ttctagctct gacatttctg atgtgaccct gggcaggtca ctgaggaagc tctctgcacc
     4321 cccatctcat tttctagaaa atcagaagaa gaaccccatg gagatgaagt gacctaaggc
     4381 cataatgtta aacgcgtgcg gccgcaggaa cccctagtga tggagttggc cactccctct
     4441 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcggc
     4501 ctcagtgagc gagcgagcgc gcagctgcct gcaggacatg tgagcaaaag gccagcaaaa
     4561 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     4621 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     4681 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     4741 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     4801 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     4861 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4921 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4981 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     5041 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     5101 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     5161 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     5221 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     5281 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     5341 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     5401 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     5461 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     5521 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     5581 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     5641 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     5701 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     5761 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     5821 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     5881 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     5941 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     6001 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     6061 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     6121 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     6181 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     6241 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     6301 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     6361 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     6421 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     6481 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     6541 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     6601 taaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     6661 attttttaac caatagaccg aaatcggcaa aatcccttat aaatcaaaag aatagcccga
     6721 gatagagttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     6781 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     6841 caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     6901 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     6961 agcgaaagga gcgggcgcta aggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     7021 cacacccgcc gcgcttaatg cgccgctaca gggc
//
