title: A novel multiplex tetra-primer ARMS-PCR for the simultaneous genotyping of six single nucleotide polymorphisms associated with female cancers.
abstract: 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.
authors.affiliation: Key Laboratory for Medical Virology, Ministry of Health-National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China.
authors.forename: Chen
authors.initials: C
authors.lastname: Zhang
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.forename: Brian Z
authors.initials: BZ
authors.lastname: Ring
authors.forename: Kai
authors.initials: K
authors.lastname: Nie
authors.forename: Mengjie
authors.initials: M
authors.lastname: Yang
authors.forename: Miao
authors.initials: M
authors.lastname: Wang
authors.forename: Hongwei
authors.initials: H
authors.lastname: Shen
authors.forename: Xiyang
authors.initials: X
authors.lastname: Wu
authors.forename: Xuejun
authors.initials: X
authors.lastname: Ma
mesh_terms: D001483:Base Sequence; D004252:DNA Mutational Analysis; D017931:DNA Primers / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D005260:Female; D005787:Gene Frequency / Q000235:genetics; D020022:Genetic Predisposition to Disease; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D008969:Molecular Sequence Data; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D009154:Mutation / Q000235:genetics*; D009369:Neoplasms / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*
chemical_list: D017931:DNA Primers