Construction and Validation of an Expression Vector for Umami Octapeptide
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Abstract:
Umami peptide is a recently identified umami substance, but its taste is controversial domestically and internationally. The possible reason for the controversy is that the method of verifying its taste is mainly based on chemical synthesis. In order to obtain natural umami peptide for the further evaluation of the umami peptide taste, the controversial umami octapeptide was used as the target peptide, gene engineering was adopted to construct an expression vector for the octapeptide, and its expression effect was verified. The DNA sequence of the octapeptide was designed according to the primary structure of the octapeptide, Lys-Gly-Asp-Glu-Glu-Ser-Leu-Ala, and codon bias of Escherichia coli BL 21 (DE3), and was cloned into pET-32a (+) to form the expression vector. Through colony polymerase chain reaction (PCR) identification of positive clones, identification of the recombinant plasmid, and the induced expression of the engineered bacteria, it was found that the target gene of the octapeptide was successfully restructured into the pET-32a vector, and the recombinant pET-32a vector-transformed E. coli could normally express the fusion protein. This method will lay a solid foundation for obtaining the natural octapeptide in the future.
