LOCUS       sequence-382349-        3840 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression vector for  a low-affinity calcium indicator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3840)
  AUTHORS   Valiente-Gabioud AA, Garteizgogeascoa Suner I, Idziak A, Fabritius 
            A, Basquin J, Angibaud J, Nagerl UV, Singh SP, Griesbeck O
  TITLE     Fluorescent sensors for imaging of interstitial calcium.
  JOURNAL   Nat Commun. 2023 Oct 5;14(1):6220. doi: 10.1038/s41467-023-41928-w.
  PUBMED    37798285
REFERENCE   2  (bases 1 to 3840)
  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"; journalName: "Nat 
            Commun."; date: "2023-10-5"; volume: "14(1):6220. doi"; pages: " 
            10.1038/s41467-023-41928-w"
COMMENT     Sequence Label: GreenT-EC.b - pRSET
FEATURES             Location/Qualifiers
     source          1..3840
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     133..152
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     386..403
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     488..507
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        488..506
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             538..560
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             580..609
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             616..648
                     /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(652..660,661..675)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     regulatory      682..691
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1487..1496
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1642..1659)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1709..1727)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1723..1770
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1867..2322
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1954..1973)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2164..2185
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2348..2452
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2453..2521,2522..3313)
                     /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(2671..2690)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(3484..3840,1..232)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
       61 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
      121 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
      181 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
      241 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
      301 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
      361 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
      421 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagga tctcgatccc
      481 gcgaaattaa tacgactcac tatagggaga ccacaacggt ttccctctag aaataatttt
      541 gtttaacttt aagaaggaga tatacatatg cggggttctc atcatcatca tcatcaccat
      601 caccatcacc gcggtatggc tagcatgact ggtggacagc aaatgggtcg ggatctgtac
      661 gacgatgacg ataaggatcc ggccaccatg gtctccaagg gggaggagga caacatgggg
      721 agcctgccag ctacccacga gctgcatatc tttggatcta ttaacggggt ggatttcgac
      781 atggttggac aaggtagcgg gaatcctaat gatgggtatg aagagcttaa ccttaaaagc
      841 actaagggcg atctgcaatt ttcaccttgg attctcgtcc cccacattgg gtatgggttc
      901 catcaatatc tcccttaccc ggatggaatg tctcccttcc aagcggctat ggtagatggt
      961 agtggttacc aagttcacag gactatgcaa ttcgaagacg gcgcctcatt gactgtgaac
     1021 taccgataca cttacgaagg atctcacata aaaggtgaag cgcaggtgaa gggcacggga
     1081 tttccagcag atgggccggt aatgactaac tcacttactg cggcagactt ggggtgggat
     1141 tcagaagagg aactttccga atgttttaga atttttgatt ttgacggtaa cggattcatt
     1201 gaccgcgaag agttcggtga cataatcaga ttgactgggg agcaacttac ggatgaggac
     1261 gtagacgaaa tttttggcga ctcagatacg gataagaatt caggccggat cgactttgat
     1321 gaatttctga aaatggtcga aaatgttcag ctcacggaca accgaacaac gaaaaagacc
     1381 tatccaaatg acaagacaat tatctcaact ttgaaatgga gctacaccac ggacaacggc
     1441 aagaggtatc ggtccaccgc tatcactact tatacatttg ctaaacccat ggcggctaac
     1501 tacctcaaga atcaacctat gtacgtgttt cgcaagactg agcttaaaca tagcaaaacc
     1561 gaattgaatt tcaaggagtg gcagaaggcg tttacggacg taatggggat ggatgagctt
     1621 tataagtgat aataggaatt cctatagtgt cacctaaatg atccggctgc taacaaagcc
     1681 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
     1741 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatctggc
     1801 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     1861 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     1921 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     1981 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     2041 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     2101 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2161 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     2221 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     2281 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
     2341 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     2401 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     2461 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     2521 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     2581 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     2641 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     2701 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     2761 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     2821 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     2881 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     2941 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     3001 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     3061 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     3121 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     3181 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     3241 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     3301 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     3361 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     3421 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     3481 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     3541 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     3601 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     3661 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     3721 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     3781 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
//
