Modified Tetra-Primer ARMS PCR optimization for detection of ADIPOQ rs267729 SNP (C>G) in type 2 diabetes patients in San Luis, Argentina population
1 Laboratory of Diabetes, Department of Biochemistry and Biological Sciences, School of Chemistry, Biochemistry and Pharmacy, National University of San Luis, San Luis, Argentina.
2 San Luis Hospital, San Luis, Argentina.
Research Article
GSC Biological and Pharmaceutical Sciences, 2020, 10(02), 138-147.
Article DOI: 10.30574/gscbps.2020.10.2.0030
Publication history:
Received on 03 February 2020; revised on 14 February 2020; accepted on 17 February 2020
Abstract:
The human Adiponectin (ADIPOQ) is an important adipocytokine that is secreted by adipocytes and plays a key role in the inflammatory response that is associated with insulin-resistant states and T2DM. The transversion of C to G (rs267729) in the promoter region of ADIPOQ gene could affect its transcription and secretion. Development of a new Modified Tetra Primer Amplification Refractory Mutation System - Polymerase Chain Reaction (MTPA-PCR) for detection of rs267729 (C>G) in the human promoter of ADIPOQ gene was sought. A MTPA-PCR for rs267729 polymorphism in a single-step PCR was carried out, and the results were confirmed by ASO-PCR technique in 25 type 2 diabetes mellitus patients and 25 controls. We have developed a sensitive single tube MTPA-PCR assay to detect rs267729 (C>G) in the human promoter of ADIPOQ gene, which allowed us to distinguish homozygous and heterozygous forms of this polymorphism successfully. Full accordance between ASO-PCR and MTPA-PCR methods for genotyping of rs267729 (C>G) polymorphism was found. MTPA-PCR could be a reliable, accurate and simple technique for genotyping SNP and different mutations. This work describes a rapid, relatively cheap, high throughput detection of C>G polymorphism in ADIPOQ that can be used in large scale clinical studies. So far, no study was done on optimization methods for genotyping rs267729 (C>G) polymorphism by MTPA-PCR. Here, we successfully adjusted and enhanced this method for recognizing rs267729 (C>G) polymorphism in the promoter of ADIPOQ gene in Argentinian population.
Keywords:
Adipocytes; Inflammatory response; Insulin-Resistant; Genotyping
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Copyright © 2020 Author(s) retain the copyright of this article. This article is published under the terms of the Creative Commons Attribution Liscense 4.0
