Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG

A potential marker, green fluorescent protein (GFP) derived from the jellyfish Aequorea victoria offer many advantages as a viability reporter, as it requires no external source of substrate nor cofactors to fluoresce but is dependent on the host cell being alive. Its use has been reported in M....

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Main Author:	Najamudin, Khairul Ezani
Format:	Thesis
Language:	English
Published:	2007
Subjects:	RS1-441 Pharmacy and materia medica
Online Access:	http://eprints.usm.my/51085/1/Khairul%20Ezani%20Najamudin_hj.pdf
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id	my-usm-ep.51085
record_format	uketd_dc
spelling	my-usm-ep.510852022-01-06T02:56:32Z Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG 2007-11 Najamudin, Khairul Ezani RS1-441 Pharmacy and materia medica A potential marker, green fluorescent protein (GFP) derived from the jellyfish Aequorea victoria offer many advantages as a viability reporter, as it requires no external source of substrate nor cofactors to fluoresce but is dependent on the host cell being alive. Its use has been reported in M. tuberculosis, but the incorporation of potentially pathogenic strains for high throughput screening would raise issues of safety. Therefore, a safe strain such as the vaccine strains M. bovis BCG if made to express an easily detectable viability * reporter marker would be useful tool as a screening assay. In this study, a synthetic version of the wild type GFP gene sequence incorporating mycobacterial codon bias, was used to explore its potential as a marker of viability in the mycobacterial vaccine strain, M. bovis BCG. The synthetic gene designated as EzyGFP was successfully constructed by using assembly polymerase chain reaction (PCR). In the second approach, a modified version of GFP, GFPuv, was also tested as an alternative candidate. To propagate these genes in Escherichia coli and then deliver these genes into M. bovis BCG, mycobacterial shuttle vectors were constructed. To create mycobacterial shuttle vectors, the mycobacterial origin of replication was excised from a source plasmid, pNMN012, and ligated into plasmids designated as pEzyGFP, pGFPuvK and pGFPuvK65 respectively which contain the GFP gene of interest. 2007-11 Thesis http://eprints.usm.my/51085/ http://eprints.usm.my/51085/1/Khairul%20Ezani%20Najamudin_hj.pdf application/pdf en public masters Perpustakaan Hamzah Sendut Pusat Pengajian Sains Kesihatan
institution	Universiti Sains Malaysia
collection	USM Institutional Repository
language	English
topic	RS1-441 Pharmacy and materia medica
spellingShingle	RS1-441 Pharmacy and materia medica Najamudin, Khairul Ezani Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG
description	A potential marker, green fluorescent protein (GFP) derived from the jellyfish Aequorea victoria offer many advantages as a viability reporter, as it requires no external source of substrate nor cofactors to fluoresce but is dependent on the host cell being alive. Its use has been reported in M. tuberculosis, but the incorporation of potentially pathogenic strains for high throughput screening would raise issues of safety. Therefore, a safe strain such as the vaccine strains M. bovis BCG if made to express an easily detectable viability * reporter marker would be useful tool as a screening assay. In this study, a synthetic version of the wild type GFP gene sequence incorporating mycobacterial codon bias, was used to explore its potential as a marker of viability in the mycobacterial vaccine strain, M. bovis BCG. The synthetic gene designated as EzyGFP was successfully constructed by using assembly polymerase chain reaction (PCR). In the second approach, a modified version of GFP, GFPuv, was also tested as an alternative candidate. To propagate these genes in Escherichia coli and then deliver these genes into M. bovis BCG, mycobacterial shuttle vectors were constructed. To create mycobacterial shuttle vectors, the mycobacterial origin of replication was excised from a source plasmid, pNMN012, and ligated into plasmids designated as pEzyGFP, pGFPuvK and pGFPuvK65 respectively which contain the GFP gene of interest.
format	Thesis
qualification_level	Master's degree
author	Najamudin, Khairul Ezani
author_facet	Najamudin, Khairul Ezani
author_sort	Najamudin, Khairul Ezani
title	Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG
title_short	Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG
title_full	Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG
title_fullStr	Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG
title_full_unstemmed	Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG
title_sort	application of aequorea victoria green fluorescent protein (gfp) for expression in mycobacterium bovis bcg
granting_institution	Perpustakaan Hamzah Sendut
granting_department	Pusat Pengajian Sains Kesihatan
publishDate	2007
url	http://eprints.usm.my/51085/1/Khairul%20Ezani%20Najamudin_hj.pdf
_version_	1747822056935784448

Application Of Aequorea Victoria Green Fluorescent Protein (Gfp) For Expression In Mycobacterium bovis BCG

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