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A Review of Optimization Strategies and the Advantages and Disadvantages of using Polymerase Chain Reaction with Confronting Two-Pair primers (PCR-CTPP) in Genomics Studies
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Zeinab Jamshidi1    , Kheirollah Yari 2 |
1- BSc Student, Biotechnology Department, Faculty of Sciences, Malayer University, Malayer, Iran
2- Assistant Professor, Medical Biology Research Center, Health Technology Institute, Kermanshah University of Medical Sciences, Kermanshah, Iran, , Khirollah.yari@yahoo.com |
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Abstract: (1514 Views) |
Background and Aim: There are several methods for genotyping single nucleotide polymorphisms (SNPs). Polymerase chain reaction with confronting two-pair primers (PCR-CTPP) is an inexpensive, quick, reliable, time saving and cost effective method that is applicable for SNPs genotyping compared to other related methods. In the PCR-CTPP technique, allele-specific products of the genes are selectively amplified by adding two pairs of four primers and ordinarily prepared PCR mixture in a single PCR tube. Four allele-specific primers are F1 (sense) and R1 (antisense) primers for one allele that R1 has an anti-sense nucleotide at the 3′ end and F2 (sense) and R2 (antisense) for the other allele that F2 has a sense nucleotide at the 3′ end. PCR-CTPP produces three products with different sizes which can make possible genotyping of SNP by gel electrophoresis. To increase the reproducibility and accuracy of the PCR-CTPP results, it is better to optimize protocol before sample genotyping. The aim of this study was to review the currently used strategies for optimum application and determination of advantages and disadvantages of the PCR-CTPP method in SNPs genotyping.
Materials and Methods: The articles related to the PCR-CTPP method were searched from databases such as ISI Web of science, Science direct, Pubmed, Google Scholar, SID, and Scopus.
Results: For more efficiency and reproducibility of the PCR-CTPP method, we should consider the specificity and similarity of melting temperatures of the designed primers by using algorithms and specific primer design softwares.
Conclusion: Strategies for optimizing the concentration ratio of internal and external primers, adding amplification additives to the PCR reaction mix, and use of the touchdown program are also recommended.
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| Keywords: PCR, PCR with confronting two-pair primers, Genomics, Single nucleotide polymorphism, Genotyping |
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Full-Text [PDF 357 kb]
(299 Downloads)
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Type of Study: Review |
Subject:
Molecular Medicine and Genetics
Received: 2021/08/24 | Accepted: 2022/01/3 | Published: 2022/12/14
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