Cloning of Bacillus subtilis phytase gene construct in Escherichia coli

Abstract

Background and Objectives: Phytase has a hydrolysis function of phytic acid, which yields inorganic phosphate. Bacillus species can produce thermostable alkaline phytase. The aim of this study was to isolate and clone a Phytase gene (Phy) from Bacillus subtilis in Escherichia coli. Materials and Methods: In this study, the extracellular PhyC gene was isolated from Bacillus subtilis Phytase C. After purification of the bands, DNA fragment of Phy gene was cloned by T/A cloning technique, and the clone was transformed into Escherichia coli. Afterward, the pGEM-Phy was transferred into E. coli Top-10 strain and the recombinants were plated on LB agar containing 100 µg/ml ampicillin. The colonization of 1171 bp of gene Phytase C was confirmed by PCR. The presence of gene-targeting in vector was confirmed with enzymatic digestion by XhoI and XbaI restriction enzymes. Results: The Phytase gene was successfully cloned in E. coli. The result of cloning of 1171 bp Phytase gene was confirmed by PCR assay. Conclusion: Our impression of this article is that several methods, such as using along with microbial, plant phytase reproduction, or low-phytic acid corn may be the better way from a single phytase.

Keywords

• Bacillus subtilis;
• Cloning;
• Escherichia coli;
• Phytase;
• Probiotics