A Unique Primer with an Inosine Chain at the 5′-Terminus Improves the Reliability of SNP Analysis Using the PCR-Amplified Product Length Polymorphism Method
نویسندگان
چکیده
Polymerase chain reaction-amplified product length polymorphism (PCR-APLP) is one of the most convenient and reliable methods for single nucleotide polymorphism (SNP) analysis. This method is based on PCR, but uses allele-specific primers containing SNP sites at the 3'-terminus of each primer. To use this method at least two allele-specific primers and one "counter-primer", which serves as a common forward or reverse primer of the allele-specific primers, are required. The allele-specific primers have SNP sites at the 3'-terminus, and another primer should have a few non-complementary flaps at the 5'-terminus to detect SNPs by determining the difference of amplicon length by PCR and subsequent electrophoresis. A major disadvantage of the addition of a non-complementary flap is the non-specific annealing of the primer with non-complementary flaps. However, a design principle for avoiding this undesired annealing has not been fully established, therefore, it is often difficult to design effective APLP primers. Here, we report allele-specific primers with an inosine chain at the 5'-terminus for PCR-APLP analysis. This unique design improves the competitiveness of allele-specific primers and the reliability of SNP analysis when using the PCR-APLP method.
منابع مشابه
Molecular analysis of the S1 gene of vaccine strains of infectious bronchitis virus using reverse transcriptase-polymerase chain reaction and restriction fragment length polymorphism
Infectious bronchitis virus (IBV) is an acute and contagious viral disease of poultry that affects different systems, including the respiratory tract in particular. IBV causes major economic losses in the poultry industry globally. Due to antigenic variation of the causative agent, control of the disease is difficult. To control the disease, many vaccines that belong to different serotypes are...
متن کاملDETECTION AND RESTRICTION ANALYSIS OF C YTOMEGALOVIRUS DNA PERSISTING IN HUMAN ATHEROSCLEROTIC PLAQUES USING POLYMERASE CHAIN REACTION
The polymerase chain reaction (PCR) as applied to detection of a foreign DNA in clinical specimens could provide a sensitive instrument for rapid detection of viral DNA persisting in tissues of patients suspected of latent infection. Human cytomegalovirus (HCMV) DNA was found in arterial plaques of patients with atherosclerotic lesions using a PCR assay with nested primer oligonucleotides ...
متن کاملBenzimidazole -Resistance in Haemonchus contortus: New PCR-RFLP Method for the Detection of Point Mutation at Codon 167 of Isotype 1 β-Tubulin Gene
BACKGROUND Due to the lack of a suitable and economic test for the analysis of the polymorphism at codon 167, we developed a new PCR-RFLP technique, based on a modified forward primer (UT-HC167 MF-primer), to identify simultaneously the SNPs at codons 167 and 200 of isotype 1 β-tubulin gene of Haemonchus contortus. METHODS There already are several safe and easy methods for identification of ...
متن کاملGenome complexity reduction for SNP genotyping analysis.
Efficient single nucleotide polymorphism (SNP) genotyping methods are necessary to accomplish many current gene discovery goals. A crucial element in large-scale SNP genotyping is the number of individual biochemical reactions that must be performed. An efficient method that can be used to simultaneously amplify a set of genetic loci across a genome with high reliability can provide a valuable ...
متن کاملAnalysis of the Genetic Diversity 12 Iranian Damask Rose (Rosa damascena Mill.) Genotypes Using Amplified Fragment Length Polymorphism Markers
In this study, the genetic diversity of 12 Iranian Damask rose (Rosa damascena Mill.) genotypes was studied using amplified fragment length polymorphism (AFLP) markers. Twelve AFLP primer combinations generated 483 polymorphic bands and showed extreme variability and genetic complexity among the studied genotypes. The AFLP analysis revealed a specific amplified fragment for the genotypes collec...
متن کامل