Cloning and Expression of Randomly Mutated Bacillus subtilisα-Amylase Genes in HB101
Joint Authors
MirMohammad Sadeghi, Hamid
Salehi, Golnaz
Moazen, Fatemeh
Rabbani, Mohammed
Rabbani, Mohammad
Source
Biotechnology Research International
Issue
Vol. 2011, Issue 2011 (31 Dec. 2011), pp.1-5, 5 p.
Publisher
Hindawi Publishing Corporation
Publication Date
2011-07-24
Country of Publication
Egypt
No. of Pages
5
Main Subjects
Information Technology and Computer Science
Abstract EN
The aim of this study was to isolate and express the randomly mutated α-amylase gene from B.
subtilis strain 168.
BS168F: 5′-gtgtcaagaatgtttgc-3′ and BS168R: 3′-gttttgttaaaagatga-5′ primers were used to amplify the amylase gene using the following cycle in error-prone PCR method: 94°C for 30 s, 40°C for 2 min, and 72°C for 2 min in 30 cycles that were followed with 72°C for 2 min as a post cycle.
E.
coli XL1 blue was used as host for plasmid construction.
Amylase enzyme activity assay was performed using continuous spectrophotometric procedures.
Results of sequencing showed that sequence was cloned from the first ATG and with the correct open reading frame.
Having confirmed the integrity of the insert, the gene was ligated into expression vector pET-15b and then further confirmed using digestion analysis.
Amylase activity showed 3 clones with higher enzymatic activity compared with the wild type.
Error-prone PCR method produced a mutated gene that provides amylase activity much higher than that of wild type.
Sequencing the mutated genes should shed light on the important region of the genes that could be manipulated in future studies.
American Psychological Association (APA)
Rabbani, Mohammed& MirMohammad Sadeghi, Hamid& Moazen, Fatemeh& Rabbani, Mohammad& Salehi, Golnaz. 2011. Cloning and Expression of Randomly Mutated Bacillus subtilisα-Amylase Genes in HB101. Biotechnology Research International،Vol. 2011, no. 2011, pp.1-5.
https://search.emarefa.net/detail/BIM-462009
Modern Language Association (MLA)
Rabbani, Mohammed…[et al.]. Cloning and Expression of Randomly Mutated Bacillus subtilisα-Amylase Genes in HB101. Biotechnology Research International No. 2011 (2011), pp.1-5.
https://search.emarefa.net/detail/BIM-462009
American Medical Association (AMA)
Rabbani, Mohammed& MirMohammad Sadeghi, Hamid& Moazen, Fatemeh& Rabbani, Mohammad& Salehi, Golnaz. Cloning and Expression of Randomly Mutated Bacillus subtilisα-Amylase Genes in HB101. Biotechnology Research International. 2011. Vol. 2011, no. 2011, pp.1-5.
https://search.emarefa.net/detail/BIM-462009
Data Type
Journal Articles
Language
English
Notes
Includes bibliographical references
Record ID
BIM-462009