LOCUS       sequence-220800-        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        8..292
                     /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     215..234
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             338..355
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             359..391
                     /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(395..403,404..418)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             449..505
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKAGHAVRAIGRLSS"
     primer_bind     1201..1220
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1244..1687
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
                     ELQDMINEVDADGDGTFDFPEFLTMMARKMNDTDSEEEIREAFRVFDKDGNGYIGAAEL
                     RHVMTDLGEKLTDEEVDEMIRVADIDGDGQVNYEEFVQMMTAK"
     primer_bind     complement(1726..1743)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1896..1982
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2074..2101
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2120..2211
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2212..2280,2281..3072)
                     /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(2430..2449)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3243..3831
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3732..3751
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3985..4002
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4017..4157
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4083..4102)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4243..4265
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4421..5299)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 ttcagagaag aaaccaattg tccatattgc atcagacatt gccgtcactg cgtcttttac
       61 tggctcttct cgctaaccaa accggtaacc ccgcttatta aaagcattct gtaacaaagc
      121 gggaccaaag ccatgacaaa aacgcgtaac aaaagtgtct ataatcacgg cagaaaagtc
      181 cacattgatt atttgcacgg cgtcacactt tgctatgcca tagcattttt atccataaga
      241 ttagcggatc ctacctgacg ctttttatcg caactctcta ctgtttctcc atacccgttt
      301 tttgggctaa caggaggaat taaccatggg gggttctcat catcatcatc atcatggtat
      361 ggctagcatg actggtggac agcaaatggg tcgggatctg tacgacgatg acgataagga
      421 tcctggctcg agcatggtcg actcttcacg tcgtaagtgg aataaggcag gtcacgcagt
      481 tagagctata ggtcggctga gctcacccgt ggtttccgag cggatgtacc ccgaggacgg
      541 cgccctgaag agcgagatca agaaggggct gaggctgaag gacggcggcc actacgccgc
      601 cgaggtcaag accacctaca aggccaagaa gcccgtgcag ctgcccggcg cctacatcgt
      661 cgacatcaag ttggacatcg tgtcccacaa cgaggactac accatcgtgg aacagtgcga
      721 acgcgccgag ggccgccact ccaccggcgg catggacgag ctgtacaagg gaggtacagg
      781 cgggagtctg gtgagcaagg gcgtggagga taacatggcc atcgtcaagg agttcatgcg
      841 cttcaagacg cacatagagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg
      901 cgagggccgc ccctacgagg cctttcagac cgctaagctg aaggtgacca agggtggccc
      961 cctgcccttc gcctgggaca tcctgtcccc tcagattatg tacggctcca aggcctacat
     1021 taagcaccca gccgacatcc ccgactactt caagctgtcc ttccccgagg gcttcaggtg
     1081 ggagcgcgtg atgaacttcg aggacggcgg cattattcac gttaaccagg actcctccct
     1141 gcaggacggc gtattcatct acaaggtgaa gctgcgcggc accaacttcc cccccgacgg
     1201 ccccgtaatg cagaagaaga ccatgggctg ggagcctacg cgtgaccaac tgactgaaga
     1261 gcagatcgca gaatttaaag aggctttctc cctatttgac aaggacgggg atgggacgat
     1321 aacaaccaag gagctgggga cggtgatgcg gtctctgggg cagaacccca cagaagcaga
     1381 gctgcaggac atgatcaatg aagtagatgc cgacggtgac ggcacattcg acttccctga
     1441 gttcctgacg atgatggcaa gaaaaatgaa tgacacagac agtgaagagg aaattagaga
     1501 agcgttccgc gtgtttgata aggacggcaa tggctacatc ggcgcagcag agcttcgcca
     1561 cgtgatgaca gaccttggag agaagttaac agatgaggag gttgatgaaa tgatcagggt
     1621 agcagacatc gatggggatg gtcaggtaaa ctacgaagag tttgtacaaa tgatgacagc
     1681 gaagtagaag cttggctgtt ttggcggatg agagaagatt ttcagcctga tacagattaa
     1741 atcagaacgc agaagcggtc tgataaaaca gaatttgcct ggcggcagta gcgcggtggt
     1801 cccacctgac cccatgccga actcagaagt gaaacgccgt agcgccgatg gtagtgtggg
     1861 gtctccccat gcgagagtag ggaactgcca ggcatcaaat aaaacgaaag gctcagtcga
     1921 aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa
     1981 atccgccggg agcggatttg aacgttgcga agcaacggcc cggagggtgg cgggcaggac
     2041 gcccgccata aactgccagg catcaaatta agcagaaggc catcctgacg gatggccttt
     2101 ttgcgtttct acaaactctt ttgtttattt ttctaaatac attcaaatat gtatccgctc
     2161 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     2221 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     2281 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     2341 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     2401 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac
     2461 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     2521 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     2581 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     2641 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     2701 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     2761 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     2821 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     2881 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     2941 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     3001 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     3061 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     3121 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     3181 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     3241 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     3301 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     3361 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     3421 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     3481 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     3541 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     3601 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     3661 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     3721 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     3781 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     3841 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     3901 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     3961 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg
     4021 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc
     4081 agtacaatct gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg
     4141 actgggtcat ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     4201 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     4261 agaggttttc accgtcatca ccgaaacgcg cgaggcagca gatcaattcg cgcgcgaagg
     4321 cgaagcggca tgcataatgt gcctgtcaaa tggacgaagc agggattctg caaaccctat
     4381 gctactccgt caagccgtca attgtctgat tcgttaccaa ttatgacaac ttgacggcta
     4441 catcattcac tttttcttca caaccggcac ggaactcgct cgggctggcc ccggtgcatt
     4501 ttttaaatac ccgcgagaaa tagagttgat cgtcaaaacc aacattgcga ccgacggtgg
     4561 cgataggcat ccgggtggtg ctcaaaagca gcttcgcctg gctgatacgt tggtcctcgc
     4621 gccagcttaa gacgctaatc cctaactgct ggcggaaaag atgtgacaga cgcgacggcg
     4681 acaagcaaac atgctgtgcg acgctggcga tatcaaaatt gctgtctgcc aggtgatcgc
     4741 tgatgtactg acaagcctcg cgtacccgat tatccatcgg tggatggagc gactcgttaa
     4801 tcgcttccat gcgccgcagt aacaattgct caagcagatt tatcgccagc agctccgaat
     4861 agcgcccttc cccttgcccg gcgttaatga tttgcccaaa caggtcgctg aaatgcggct
     4921 ggtgcgcttc atccgggcga aagaaccccg tattggcaaa tattgacggc cagttaagcc
     4981 attcatgcca gtaggcgcgc ggacgaaagt aaacccactg gtgataccat tcgcgagcct
     5041 ccggatgacg accgtagtga tgaatctctc ctggcgggaa cagcaaaata tcacccggtc
     5101 ggcaaacaaa ttctcgtccc tgatttttca ccaccccctg accgcgaatg gtgagattga
     5161 gaatataacc tttcattccc agcggtcggt cgataaaaaa atcgagataa ccgttggcct
     5221 caatcggcgt taaacccgcc accagatggg cattaaacga gtatcccggc agcaggggat
     5281 cattttgcgc ttcagccata cttttcatac tcccgcca
//
