A Novel Multiplex Tetra-Primer ARMS-PCR for the Simultaneous Genotyping of Six Single Nucleotide Polymorphisms Associated with Female CancersReportar como inadecuado




A Novel Multiplex Tetra-Primer ARMS-PCR for the Simultaneous Genotyping of Six Single Nucleotide Polymorphisms Associated with Female Cancers - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

Background

The tetra-primer amplification refractory mutation system PCR T-ARMS-PCR is a fast and economical means of assaying SNP-s, requiring only PCR amplification and subsequent electrophoresis for the determination of genotypes. To improve the throughput and efficiency of T-ARMS-PCR, we combined T-ARMS-PCR with a chimeric primer-based temperature switch PCR TSP strategy, and used capillary electrophoresis CE for amplicon separation and identification. We assessed this process in the simultaneous genotyping of four breast cancer–and two cervical cancer risk–related SNPs.

Methods

A total of 24 T-ARMS-PCR primers, each 5′-tagged with a universal sequence and a pair of universal primers, were pooled together to amplify the 12 target alleles of 6 SNPs in 186 control female blood samples. Direct sequencing of all samples was also performed to assess the accuracy of this method.

Results

Of the 186 samples, as many as 11 amplicons can be produced in one single PCR and separated by CE. Genotyping results of the multiplex T-ARMS-PCR were in complete agreement with direct sequencing of all samples.

Conclusions

This novel multiplex T-ARMS-PCR method is the first reported method allowing one to genotype six SNPs in a single reaction with no post-PCR treatment other than electrophoresis. This method is reliable, fast, and easy to perform.



Autor: Chen Zhang , Ying Liu , Brian Z. Ring, Kai Nie, Mengjie Yang, Miao Wang, Hongwei Shen, Xiyang Wu , Xuejun Ma

Fuente: http://plos.srce.hr/



DESCARGAR PDF




Documentos relacionados