Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity

The p53 therapy based approach is a good candidate for treatment of TP53-defect cancer. However, this treatment is unsuitable for some cancer cases especially for that caused by dominant-negative activity of the mutant p53 protein. Thus, more effective treatment is needed to overcome this problem an...

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Main Authors:	Bakhtiar, M. Taher, Solachuddin Jauhari , Arief Ichwan
Format:	Conference or Workshop Item
Language:	English
Published:	2012
Subjects:	RS Pharmacy and materia medica
Online Access:	http://irep.iium.edu.my/22804/ http://irep.iium.edu.my/22804/ http://irep.iium.edu.my/22804/1/zafirah_ICBS_abstract.pdf

id	iium-22804
recordtype	eprints
spelling	iium-228042014-12-08T07:35:30Z http://irep.iium.edu.my/22804/ Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity Bakhtiar, M. Taher Solachuddin Jauhari , Arief Ichwan RS Pharmacy and materia medica The p53 therapy based approach is a good candidate for treatment of TP53-defect cancer. However, this treatment is unsuitable for some cancer cases especially for that caused by dominant-negative activity of the mutant p53 protein. Thus, more effective treatment is needed to overcome this problem and DNA vaccination may be the suitable candidate where mutant p53 may become a suitable candidate as an antigen for the DNA vaccination strategy. Therefore, the research aims to produce mutant TP53R248Q through PCR site–directed mutagenesis and to confirm its tumor suppression ability. The mutation of R248Q was generated via Polymerase Chain Reaction (PCR) site-directed mutagenesis to pCMVp53 plasmid. Following that, these constructed TP53R248Q was transfected into the TP53-null H1299 cell lines in order to investigate its tumor suppression ability, by studying its phenotype and genotype expression. Phenotype study was conducted by using colony formation assay while quantification of TP53 downstream target gene, p21waf1 was conducted for the genotype study. The transfection with exogenous TP53R248Q resulted in massive colony proliferation and downregulation of p21waf1, thus confirmed that the generated mutant TP53 has lost its ability to restrain cells growth. These data therefore confirmed that the PCR site-directed mutagenesis technique has been successfully carried out to induce the desired mutation in the TP53 gene. Thus, this technique may become an interesting option to generate novel recombinant proteins, especially in the development of specifically designed DNA vaccine as a gene therapy in the future. 2012-02-27 Conference or Workshop Item NonPeerReviewed application/pdf en http://irep.iium.edu.my/22804/1/zafirah_ICBS_abstract.pdf Bakhtiar, M. Taher and Solachuddin Jauhari , Arief Ichwan (2012) Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity. In: International Conference in Biomedical Science 2012, 27-28 February, Bandung. http://www.sith.itb.ac.id/biomedical_conference/getToBandung.php
repository_type	Digital Repository
institution_category	Local University
institution	International Islamic University Malaysia
building	IIUM Repository
collection	Online Access
language	English
topic	RS Pharmacy and materia medica
spellingShingle	RS Pharmacy and materia medica Bakhtiar, M. Taher Solachuddin Jauhari , Arief Ichwan Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity
description	The p53 therapy based approach is a good candidate for treatment of TP53-defect cancer. However, this treatment is unsuitable for some cancer cases especially for that caused by dominant-negative activity of the mutant p53 protein. Thus, more effective treatment is needed to overcome this problem and DNA vaccination may be the suitable candidate where mutant p53 may become a suitable candidate as an antigen for the DNA vaccination strategy. Therefore, the research aims to produce mutant TP53R248Q through PCR site–directed mutagenesis and to confirm its tumor suppression ability. The mutation of R248Q was generated via Polymerase Chain Reaction (PCR) site-directed mutagenesis to pCMVp53 plasmid. Following that, these constructed TP53R248Q was transfected into the TP53-null H1299 cell lines in order to investigate its tumor suppression ability, by studying its phenotype and genotype expression. Phenotype study was conducted by using colony formation assay while quantification of TP53 downstream target gene, p21waf1 was conducted for the genotype study. The transfection with exogenous TP53R248Q resulted in massive colony proliferation and downregulation of p21waf1, thus confirmed that the generated mutant TP53 has lost its ability to restrain cells growth. These data therefore confirmed that the PCR site-directed mutagenesis technique has been successfully carried out to induce the desired mutation in the TP53 gene. Thus, this technique may become an interesting option to generate novel recombinant proteins, especially in the development of specifically designed DNA vaccine as a gene therapy in the future.
format	Conference or Workshop Item
author	Bakhtiar, M. Taher Solachuddin Jauhari , Arief Ichwan
author_facet	Bakhtiar, M. Taher Solachuddin Jauhari , Arief Ichwan
author_sort	Bakhtiar, M. Taher
title	Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity
title_short	Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity
title_full	Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity
title_fullStr	Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity
title_full_unstemmed	Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity
title_sort	site-directed mutagenesis for the production of mutant tp53 gene and analysis of its tumor suppressor activity
publishDate	2012
url	http://irep.iium.edu.my/22804/ http://irep.iium.edu.my/22804/ http://irep.iium.edu.my/22804/1/zafirah_ICBS_abstract.pdf
first_indexed	2023-09-18T20:34:39Z
last_indexed	2023-09-18T20:34:39Z
_version_	1777408965235179520

Site-directed mutagenesis for the production of mutant TP53 gene and analysis of its tumor suppressor activity

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