LOCUS       sequence-99657-e        2081 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Protein expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 2081)
  AUTHORS   Goulas T, Cuppari A, Garcia-Castellanos R, Snipas S, Glockshuber R, 
            Arolas JL, Gomis-Ruth FX
  TITLE     The pCri System: A Vector Collection for Recombinant Protein 
            Expression and Purification.
  JOURNAL   PLoS One. 2014 Nov 11;9(11):e112643. doi: 
            10.1371/journal.pone.0112643. eCollection 2014.
  PUBMED    25386923
REFERENCE   2  (bases 1 to 2081)
  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: "PLoS One.";
            date: "2014-11-11"; volume: "9(11):e112643. doi"; pages: " 
            10.1371/journal.pone.0112643. eCollection 2014"
COMMENT     Sequence Label: pCri-1b
FEATURES             Location/Qualifiers
     source          1..2081
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     RBS             33..55
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             68..85
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             89..1189
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1163..1186
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1214..1234
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(1250..1252,1253..1966)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1292..1313)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1553..1572)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1900..1921
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2019..2036
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 gtttggacgg aacaattccc ctctagaata attttgttta actttaagaa ggagatatac
       61 catgaaacat caccatcacc atcaccccat gaaaatcgaa gaaggtaaac tggtaatctg
      121 gattaacggc gataaaggct ataacggtct cgctgaagtc ggtaagaaat tcgagaaaga
      181 taccggaatt aaagtcaccg ttgagcatcc ggataaactg gaagagaaat tcccacaggt
      241 tgcggcaact ggcgatggcc ctgacattat cttctgggca cacgaccgct ttggtggcta
      301 cgctcaatct ggcctgttgg ctgaaatcac cccggacaaa gcgttccagg acaagctgta
      361 tccgtttacc tgggatgccg tacgttacaa cggcaagctg attgcttacc cgatcgctgt
      421 tgaagcgtta tcgctgattt ataacaaaga tctgctgccg aacccgccaa aaacctggga
      481 agagatcccg gcgctggata aagaactgaa agcgaaaggt aagagcgcgc tgatgttcaa
      541 cctgcaagaa ccgtacttca cctggccgct gattgctgct gacgggggtt atgcgttcaa
      601 gtatgaaaac ggcaagtacg acattaaaga cgtgggcgtg gataacgctg gcgcgaaagc
      661 gggtctgacc ttcctggttg acctgattaa aaacaaacac atgaatgcag acaccgatta
      721 ctccatcgca gaagctgcct ttaataaagg cgaaacagcg atgaccatca acggcccgtg
      781 ggcatggtcc aacatcgaca ccagcaaagt gaattatggt gtaacggtac tgccgacctt
      841 caagggtcaa ccatccaaac cgttcgttgg cgtgctgagc gcaggtatta acgccgccag
      901 tccgaacaaa gagctggcaa aagagttcct cgaaaactat ctgctgactg atgaaggtct
      961 ggaagcggtt aataaagaca aaccgctggg tgccgtagcg ctgaagtctt acgaggaaga
     1021 gttggcgaaa gatccacgta ttgccgccac tatggaaaac gcccagaaag gtgaaatcat
     1081 gccgaacatc ccgcagatgt ccgctttctg gtatgccgtg cgtactgcgg tgatcaacgc
     1141 cgccagcggt cgtcagactg tcgatgaagc cctgaaagac gcgcagacta attcgggatc
     1201 tggcagtggt tctgagaatc tttattttca gggccatatg gctagcaaag tgagcaaggg
     1261 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     1321 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     1381 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt
     1441 cggctacggc ctgcagtgct tcgcccgcta ccccgaccac atgaagcagc acgacttctt
     1501 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     1561 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     1621 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     1681 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     1741 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     1801 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca
     1861 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     1921 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtaaggta ccggatccga
     1981 attcgagctc cgtcgacaag cttgcggccg cactcgagca ccaccaccac caccactgag
     2041 atccggctgc taacaaagcc cgaaagaaag ctgaatggcc c
//
