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Identification of Important Polymorphic Sites on MSH2 Gene in Colorectal Cancer Samples Using High Resolution Melting Point Analysis
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M Nemati1    , AH Angji 2, S Aryan1 , A Bahari3 |
1- Department of Biochemical and molecular Sciences, Kharazmi University, Tehran, Iran,
2- Department of Biochemical and molecular Sciences, Kharazmi University, Tehran, Iran, , ershad110@yahoo.com
3- Department of Biotechnology, New Biotechnology Research Center, Zanjan University, Zanjan, Iran |
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Abstract: (3087 Views) |
Abstract
Background and Aim: Colorectal cancer (CRC) is the second most common cancer in developed countries. More than 10% of CRCs are inherited and include HNPCC and FAP ligation syndrome. The MSH2 gene is located on chromosome 2 (p21) and consists of 16 exons. MSH2 is a protein that plays a role in restorative MMR after DNA replication. The MSH2 protein is attached to MSH6 or MSH3 and forms MutSα and MutSβ complexes that identify small and large deletion / insertion abnormalities, respectively. The main objective of the present research was to evaluate HRMA's ability and effectiveness in detecting tri-nucleotide deletion mutations and determine the frequency of these mutations in homozygous and heterozygous states in colon cancer samples compared to control group.
Methods: In the present study, a case control was carried out in 2016-2017 to detect the mutations of the three nucleotide deletions of GTG in the position of exon 12 of MSH2 gene. For this purpose, 50 samples of colorectal cancer with 50 healthy samples were studied. At first, genomic DNA was extracted from paraffin tissue samples and then examined by HRMA technique and direct sequencing of the triple nucleotide deletion mutant GTG. The controls used included a sequence of 96 bat games in wild type and 93 bp in the mutant mode. Equal portions of these two were used for the heterozygous state of this site. Finally, the number of samples in each group was compared in one way ANOVA and LSD mean comparison.
Results: The number of heterozygous samples for the site in colon cancer samples was significantly higher than that of homozygous consecutive homozygos. In general, the results of the present study indicated that the HRMA technique had the ability to separate and remove the pairs of homozygos and heterozygotes according to their melting temperature (Tm)
Conclusion: According to the results of this study, the frequency of removal of the three nucleotide GTGs in cancerous samples was significantly higher than healthy ones.
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| Keywords: Colorectal Cancer, HNPCC, HRMA, MSH2 Gene |
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Full-Text [PDF 2640 kb]
(692 Downloads)
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Type of Study: Research |
Subject:
Biochemistry
Received: 2017/08/27 | Accepted: 2018/08/17 | Published: 2018/10/2
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