LOCUS       sequence-109597-         825 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expression of the Azure (blue) spectral variant of eGFP in bacteria 
            and in mammalian cells. Could be used as a reporter for quantifying 
            gene targeting and recombineering in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 825)
  AUTHORS   Valledor M, Hu Q, Schiller P, Myers RS
  TITLE     Fluorescent protein engineering by in vivo site-directed 
            mutagenesis.
  JOURNAL   IUBMB Life. 2012 Aug;64(8):684-9. doi: 10.1002/iub.1041. Epub 2012 
            May 28.
  PUBMED    22639380
REFERENCE   2  (bases 1 to 825)
  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: "10.1002/iub.1041"; 
            journalName: "IUBMB Life"; date: "2012-08"; volume: "64"; issue: 
            "8"; pages: "684-9"
COMMENT     Sequence Label: pDual-eGFP(H66)
FEATURES             Location/Qualifiers
     source          1..825
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     69..88
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        69..87
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    88..112
                     /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             127..149
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             168..185
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             195..212
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             216..248
                     /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"
     regulatory      267..276
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(273..275,276..278,279..791)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTHGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     primer_bind     complement(318..339)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(579..598)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
ORIGIN
        1 catttcaggt gtcgtgagga attctgcagt cgacggtacc gcgggcccag atctcgatcc
       61 cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga
      121 aataattttg tttaacttta agaaggagat ataccatggg cagcagccat catcatcatc
      181 atcacagcag cggcctggtg ccgcgcggca gccatatggc tagcatgact ggtggacagc
      241 aaatgggtcg cggggatcca ccggtcgcca ccatggtgag caagggcgag gagctgttca
      301 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
      361 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
      421 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc cacggcgtgc
      481 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
      541 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
      601 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
      661 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
      721 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
      781 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagc
//
