LOCUS       sequence-215156-        5318 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of red fluorescent Ca2+ sensor CAR-GECO1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5318)
  AUTHORS   Molina RS, Qian Y, Wu J, Shen Y, Campbell RE, Drobizhev M, Hughes TE
  TITLE     Understanding the Fluorescence Change in Red Genetically Encoded 
            Calcium Ion Indicators.
  JOURNAL   Biophys J. 2019 May 21;116(10):1873-1886. doi: 
            10.1016/j.bpj.2019.04.007. Epub 2019 Apr 13.
  PUBMED    31054773
REFERENCE   2  (bases 1 to 5318)
  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: 
            "10.1016/j.bpj.2019.04.007"; journalName: "Biophys J"; date: 
            "2019-05-21- 21"; volume: "116"; issue: "10"; pages: "1873-1886"
COMMENT     Sequence Label: pBAD-CAR-GECO1
FEATURES             Location/Qualifiers
     source          1..5318
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             331..348
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             352..384
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(388..396,397..411)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             442..498
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKAGHAVRAIGRLSS"
     primer_bind     1194..1213
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1237..1680
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
                     ELQDMINEVDADGDGTFDFPEFLTMMARKMNDTDSEEEIREAFRVFDKDGNGYIGAAEL
                     RHVMTDLGEKLTDEEVDEMIRVADIDGDGQVNYEEFVQMMTAK"
     primer_bind     complement(1719..1736)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1889..1975
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2067..2094
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2113..2204
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2205..2273,2274..3065)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2423..2442)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3236..3824
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3725..3744
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3978..3995
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4010..4150
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4076..4095)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4236..4258
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4414..5292)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
      301 taacaggagg aattaaccat ggggggttct catcatcatc atcatcatgg tatggctagc
      361 atgactggtg gacagcaaat gggtcgggat ctgtacgacg atgacgataa ggatcctggc
      421 tcgagcatgg tcgactcttc acgtcgtaag tggaataagg caggtcacgc agttagagct
      481 ataggtcggc tgagctcacc cgtggtttcc gagcggatgt accccgagga cggcgccctg
      541 aagagcgaga tcaagaaggg gctgaggctg aaggacggcg gccactacgc cgccgaggtc
      601 aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacat cgtcgacatc
      661 aagttggaca tcgtgtccca caacgaggac tacaccatcg tggaacagtg cgaacgcgcc
      721 gagggccgcc actccaccgg cggcatggac gagctgtaca agggaggtac aggcgggagt
      781 ctggtgagca agggcgtgga ggataacatg gccatcgtca aggagttcat gcgcttcaag
      841 acgcacatag agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc
      901 cgcccctacg aggcctttca gaccgctaag ctgaaggtga ccaagggtgg ccccctgccc
      961 ttcgcctggg acatcctgtc ccctcagatt atgtacggct ccaaggccta cattaagcac
     1021 ccagccgaca tccccgacta cttcaagctg tccttccccg agggcttcag gtgggagcgc
     1081 gtgatgaact tcgaggacgg cggcattatt cacgttaacc aggactcctc cctgcaggac
     1141 ggcgtattca tctacaaggt gaagctgcgc ggcaccaact tcccccccga cggccccgta
     1201 atgcagaaga agaccatggg ctgggagcct acgcgtgacc aactgactga agagcagatc
     1261 gcagaattta aagaggcttt ctccctattt gacaaggacg gggatgggac gataacaacc
     1321 aaggagctgg ggacggtgat gcggtctctg gggcagaacc ccacagaagc agagctgcag
     1381 gacatgatca atgaagtaga tgccgacggt gacggcacat tcgacttccc tgagttcctg
     1441 acgatgatgg caagaaaaat gaatgacaca gacagtgaag aggaaattag agaagcgttc
     1501 cgcgtgtttg ataaggacgg caatggctac atcggcgcag cagagcttcg ccacgtgatg
     1561 acagaccttg gagagaagtt aacagatgag gaggttgatg aaatgatcag ggtagcagac
     1621 atcgatgggg atggtcaggt aaactacgaa gagtttgtac aaatgatgac agcgaagtag
     1681 aagcttggct gttttggcgg atgagagaag attttcagcc tgatacagat taaatcagaa
     1741 cgcagaagcg gtctgataaa acagaatttg cctggcggca gtagcgcggt ggtcccacct
     1801 gaccccatgc cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt ggggtctccc
     1861 catgcgagag tagggaactg ccaggcatca aataaaacga aaggctcagt cgaaagactg
     1921 ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc
     1981 gggagcggat ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag gacgcccgcc
     2041 ataaactgcc aggcatcaaa ttaagcagaa ggccatcctg acggatggcc tttttgcgtt
     2101 tctacaaact cttttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     2161 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     2221 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     2281 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     2341 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     2401 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc
     2461 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     2521 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     2581 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     2641 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     2701 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     2761 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     2821 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     2881 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     2941 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     3001 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     3061 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     3121 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     3181 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     3241 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     3301 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     3361 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
     3421 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     3481 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     3541 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     3601 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     3661 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     3721 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     3781 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     3841 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     3901 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     3961 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt
     4021 attttctcct tacgcatctg tgcggtattt cacaccgcat atggtgcact ctcagtacaa
     4081 tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt
     4141 catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     4201 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     4261 ttcaccgtca tcaccgaaac gcgcgaggca gcagatcaat tcgcgcgcga aggcgaagcg
     4321 gcatgcataa tgtgcctgtc aaatggacga agcagggatt ctgcaaaccc tatgctactc
     4381 cgtcaagccg tcaattgtct gattcgttac caattatgac aacttgacgg ctacatcatt
     4441 cactttttct tcacaaccgg cacggaactc gctcgggctg gccccggtgc attttttaaa
     4501 tacccgcgag aaatagagtt gatcgtcaaa accaacattg cgaccgacgg tggcgatagg
     4561 catccgggtg gtgctcaaaa gcagcttcgc ctggctgata cgttggtcct cgcgccagct
     4621 taagacgcta atccctaact gctggcggaa aagatgtgac agacgcgacg gcgacaagca
     4681 aacatgctgt gcgacgctgg cgatatcaaa attgctgtct gccaggtgat cgctgatgta
     4741 ctgacaagcc tcgcgtaccc gattatccat cggtggatgg agcgactcgt taatcgcttc
     4801 catgcgccgc agtaacaatt gctcaagcag atttatcgcc agcagctccg aatagcgccc
     4861 ttccccttgc ccggcgttaa tgatttgccc aaacaggtcg ctgaaatgcg gctggtgcgc
     4921 ttcatccggg cgaaagaacc ccgtattggc aaatattgac ggccagttaa gccattcatg
     4981 ccagtaggcg cgcggacgaa agtaaaccca ctggtgatac cattcgcgag cctccggatg
     5041 acgaccgtag tgatgaatct ctcctggcgg gaacagcaaa atatcacccg gtcggcaaac
     5101 aaattctcgt ccctgatttt tcaccacccc ctgaccgcga atggtgagat tgagaatata
     5161 acctttcatt cccagcggtc ggtcgataaa aaaatcgaga taaccgttgg cctcaatcgg
     5221 cgttaaaccc gccaccagat gggcattaaa cgagtatccc ggcagcaggg gatcattttg
     5281 cgcttcagcc atacttttca tactcccgcc attcagag
//
