Gene modification and expression of α cold-adapted amylase in Aeromonas
[Objective]In order to obtain the low-temperature amylase gene and its related functions,and better apply it in industrial production.[Methods]Cloning of the low temperature ɑ-amylase gene from the starting strain LA7 to BL-21(DE3).Other known low temperature ɑ-amylase gene was homology anal-ysis,design specific primers,and C13 fragment was obtained by PCR amplification.Then clone it to pMAL-2X,Convert it to BL-21(DE3),and screening blue and white spots.The ɑ-amylase gene was verified by PCR and double enzyme digestion of EcoRⅠand Hind Ⅲ,the highly expressed bioengineering strain pMAL-2X-C13was obtained.The site directed mutagenic primers was design,take pMAL-2X-C13 as the tem-plate,three mutant strains of low temperature amylase gene C19,C29 and C43 were obtained.[Results]SDS-PAGE test result display that the molecular size of the protein is 114 kD and the protein expression of mutant strain C19 was higher than that of strain pMAL-2X-C13.[Conclusion]The protein expressed of mutant strain C19 more than the original strain pMAL-2X-C13.It provides a scientific and theoretical basis for the future application of the engineering bacteria in industrial production.
万方数据
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