LOCUS       sequence-460020-        1144 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Bacterial Expression of recombinant His-tagged E1GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1144)
  AUTHORS   De Lorenzi V, Ghignoli S, Bernardi M, Matteoli G, Ferri G, Storti B,
            Bizzarri R, Cardarelli F
  TITLE     Measuring pH in insulin secretory granules by phasor-based 
            fluorescence lifetime imaging of a genetically encoded sensor.
  JOURNAL   Commun Biol. 2025 Feb 25;8(1):304. doi: 10.1038/s42003-025-07758-w.
  PUBMED    40000885
REFERENCE   2  (bases 1 to 1144)
  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: "Commun 
            Biol."; date: "2025-02-25"; volume: "8(1):304. doi"; pages: " 
            10.1038/s42003-025-07758-w"
COMMENT     Sequence Label: pET-His-E1GFP
FEATURES             Location/Qualifiers
     source          1..1144
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     238..257
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     341..360
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        341..359
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    360..384
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             399..421
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             440..457
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             467..484
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             join(488..490,491..493,494..1006)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     primer_bind     complement(533..554)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(794..813)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
ORIGIN
        1 cnnnnaaagg ntttgcnnca ttcgatgnnn nccgggatcn cgacgcnnnc ccttatgcga
       61 ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag
      121 gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat
      181 acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt
      241 gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat
      301 gcgtccggcg tagaggatcg agatctcgat cccgcgaaat taatacgact cactataggg
      361 gaattgtgag cggataacaa ttcccctcta gaaataattt tgtttaactt taagaaggag
      421 atataccatg ggcagcagcc atcatcatca tcatcacagc agcggcctgg tgccgcgcgg
      481 cagccatatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
      541 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
      601 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
      661 gcccaccctc gtgaccaccc tgagctacgg cgtgcagtgc ttcagccgct accccgacca
      721 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
      781 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
      841 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
      901 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
      961 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     1021 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     1081 caaccactac ctgagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     1141 catg
//
