Optimization of recombinant amylase expression using response surface methodology (RSM)

The Anoxybaccilus DT3-1 is a newly found bacterium that is able to express amylase. The gene that encodes the amylase was recently cloned and expressed in E. coli system. However, the expression level was far too low to be used. The main objective of this study is to enhance the recombinant amylase...

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Main Author:	Muniandy, Kavitha
Format:	Thesis
Language:	English
Published:	2010
Subjects:	QA Mathematics T Technology (General)
Online Access:	http://eprints.utm.my/id/eprint/12710/6/KavithaMuniandyMFBB2010.pdf
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id	my-utm-ep.12710
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spelling	my-utm-ep.127102017-09-13T04:22:10Z Optimization of recombinant amylase expression using response surface methodology (RSM) 2010 Muniandy, Kavitha QA Mathematics T Technology (General) The Anoxybaccilus DT3-1 is a newly found bacterium that is able to express amylase. The gene that encodes the amylase was recently cloned and expressed in E. coli system. However, the expression level was far too low to be used. The main objective of this study is to enhance the recombinant amylase expression level using pET-22b vector. Another objective of this study is to determine the end product release by the reaction of this amylase. The media optimization was carried out with five different media i.e. LB, TB, SB, CDM 1 and CDM 2. Medium LB was found to be the best medium to support the cell growth and amylase production (72 U/ml). Relevant factors such as the inducer (IPTG) concentration, yeast extract concentration and induction time (OD600nm) were optimized through two Response Surface Methodology (RSM) methods, which were the Two-level factorial and Central Composite Design (CCD). After the final optimization using CCD, 83 U/ml of amylase activity was obtained with the optimal condition of 0.007 mM IPTG, 0.3% of yeast extract and induction should be done when the cells optical density was at 1.52. Upon achieving the optimal conditions, the end products were determined using High Performance Liquid Chromatography (HPLC). The amylase was able to degrade various starches like rice, corn, wheat and soluble starch and produced a wide variety of oligosaccharides such as the glucose, maltose and isomers of maltose. 2010 Thesis http://eprints.utm.my/id/eprint/12710/ http://eprints.utm.my/id/eprint/12710/6/KavithaMuniandyMFBB2010.pdf application/pdf en public masters Universiti Teknologi Malaysia, Faculty of Biosciences and Bioengineering Faculty of Biosciences and Bioengineering
institution	Universiti Teknologi Malaysia
collection	UTM Institutional Repository
language	English
topic	QA Mathematics T Technology (General)
spellingShingle	QA Mathematics T Technology (General) Muniandy, Kavitha Optimization of recombinant amylase expression using response surface methodology (RSM)
description	The Anoxybaccilus DT3-1 is a newly found bacterium that is able to express amylase. The gene that encodes the amylase was recently cloned and expressed in E. coli system. However, the expression level was far too low to be used. The main objective of this study is to enhance the recombinant amylase expression level using pET-22b vector. Another objective of this study is to determine the end product release by the reaction of this amylase. The media optimization was carried out with five different media i.e. LB, TB, SB, CDM 1 and CDM 2. Medium LB was found to be the best medium to support the cell growth and amylase production (72 U/ml). Relevant factors such as the inducer (IPTG) concentration, yeast extract concentration and induction time (OD600nm) were optimized through two Response Surface Methodology (RSM) methods, which were the Two-level factorial and Central Composite Design (CCD). After the final optimization using CCD, 83 U/ml of amylase activity was obtained with the optimal condition of 0.007 mM IPTG, 0.3% of yeast extract and induction should be done when the cells optical density was at 1.52. Upon achieving the optimal conditions, the end products were determined using High Performance Liquid Chromatography (HPLC). The amylase was able to degrade various starches like rice, corn, wheat and soluble starch and produced a wide variety of oligosaccharides such as the glucose, maltose and isomers of maltose.
format	Thesis
qualification_level	Master's degree
author	Muniandy, Kavitha
author_facet	Muniandy, Kavitha
author_sort	Muniandy, Kavitha
title	Optimization of recombinant amylase expression using response surface methodology (RSM)
title_short	Optimization of recombinant amylase expression using response surface methodology (RSM)
title_full	Optimization of recombinant amylase expression using response surface methodology (RSM)
title_fullStr	Optimization of recombinant amylase expression using response surface methodology (RSM)
title_full_unstemmed	Optimization of recombinant amylase expression using response surface methodology (RSM)
title_sort	optimization of recombinant amylase expression using response surface methodology (rsm)
granting_institution	Universiti Teknologi Malaysia, Faculty of Biosciences and Bioengineering
granting_department	Faculty of Biosciences and Bioengineering
publishDate	2010
url	http://eprints.utm.my/id/eprint/12710/6/KavithaMuniandyMFBB2010.pdf
_version_	1747814949992792064

Optimization of recombinant amylase expression using response surface methodology (RSM)

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