Cloning of α-Amylase enzyme producing gene into nonα-amylase producing organism and its Expression

 
 
International Journal of Biotech Trends and Technology (IJBTT)
 
© 2014 by IJBTT Journal
Volume - 4 Issue - 1                          
Year of Publication : 2014
Authors :Parvati Saiyam , Kuldeep Diwedi , Sujeet K. Singh , Shruti Shukla , Madhulika Singh
  10.14445/22490183/IJBTT-V4P601

"Cloning of α-Amylase enzyme producing gene into nonα-amylase producing organism and its Expression",International Journal of Biotech Trends and Technology (IJBTT), V4(1):1-3 Jan - Mar 2014. Published by Seventh Sense Research Group.

Abstract

In this work, the gene of α-amylase from B. subtilis was cloned and expressed in Non-amylase produsing E.coli. B. subtilis was isolated by performing various biochemical tests. The organism producing α-amylase enzyme was successfully isolated. The isolation of genomic DNA followed by amplification of Gene Of Interest using α-amylase primer and annealing temperature 56°C was done using PCR technique. The amplified product was eluted from electrophoresis gel and subjected to restriction digestion (EcoRI) and then ligation with the vector (Plasmid DNA pUC18). A competent cell was prepared using chemical method (CaCl2). The ligation mixture was allowed to react with the competent cells in order to produce transformed cells producing α-amylase enzyme. The mixture was spread over the starch plates. The successfully transformed E.coli cells were observed over the Starch agar plate.

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Keywords
Bacillus subtilis, E.coli, PCR, pUC18, EcoRI, etc.