Cloning of Bacillus subtilis phytase gene construct in Escherichia coli
Related Experiment Video
Updated: Oct 10, 2025
Subcloning Plus Insertion SPI - A Novel Recombineering Method for the Rapid Construction of Gene Targeting Vectors
Published on: January 8, 2015
Mahdiyar Iravani Saadi1, Abbas Doosti2, Heeva Jalali3
1Hematology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.
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.
