Production of recombinant thermophilic alpha-amylase active at low pH

摘要

Production of thermophilic alpha -amylase is new and comprises expressing Thermococcus hydrothermalis alpha -amylase gene in Escherichia coli. Production of thermophilic alpha -amylase comprises: (a) preparing an oligonucleotide probe specific for the alpha -amylase gene of a Thermococcus hydrothermalis strain (CNCM I-1319), the probe being SA221-Dig (digoxigenin-labeled sequence of 221 nucleotides given in the specification); (b) digesting chromosomal DNA of the T. hydrothermalis strain with EcoRI; (c) detecting the alpha -amylase gene by hybridization with SA221-Dig; (d) recovering DNA fragments by electrophoresis and ligating them into plasmid pKS-; (e) transforming Escherichia coli DH5 alpha with the ligation mixture by the calcium chloride method; and (f) extracting plasmids from the bacteria and recovering plasmid pEAMY101 containing the alpha -amylase gene. An Independent claim is also included for an alpha -amylase enzyme produced by the process.