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Cloning and Characterization of Lipase Gene from Soil Metagenome

Jithesh, K. (2010) Cloning and Characterization of Lipase Gene from Soil Metagenome. [Student Project Report]

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Abstract

This Dissertation / Report is the outcome of investigation carried out by the creator(s) / author(s) at the department/division of Central Food Technological Research Institute (CFTRI), Mysore mentioned below in this page.

Item Type: Student Project Report
Additional Information: DNA was extracted from the soil samples collected from various regions. Some of the extracted DNA samples were contaminated with the humic substances present in the soil. Therefore pretreatment was given to soil samples and it was observed that Aluminium sulphate precipitation method was best to remove the contaminants. The soil DNA was amplified with Lipase specific primers and an average size of 1500 bp amplified products were obtained. Amplicon was cloned and transformed into Escherichia coli JM 109 using TA cloning kit. Out of about 2000 transformed colonies, 41 were randomly selected for further studies. The recombinant Escherichia coli were screened for the lipolytic activity on chromogenic plates which contain phenol red as pH indicator dye. Out of 41, 15 transformants showed lipolytic activity on the chromogenic plates. Lipolytic activity were further confirmed on Rhodamine B agar Plates. Two of the 15 transformants showed highest lipolytic activity and these recombinants were used for the production and characterization of Lipase enzyme. From the studies it is noted that Lipase production was maximum at a temperature of 400C and an optimum pH of 8.0. At pH 8.0 the enzyme showed an activity of 5.2 U/mL and at 400C, activity of 3.65 U/mL was observed. Above and below these temperature and pH, the Lipase production was decreased.
Uncontrolled Keywords: Microorganisms, lipases, lipolytic enzymes, metagenomics,
Subjects: 600 Technology > 08 Food technology > 16 Nutritive value > 05 Enzymes
500 Natural Sciences and Mathematics > 07 Life Sciences > 04 Microbiology
Divisions: Food Microbiology
Depositing User: Food Sci. & Technol. Information Services
Date Deposited: 30 Nov 2010 05:49
Last Modified: 28 Dec 2011 10:19
URI: http://ir.cftri.com/id/eprint/9803

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