title: Single nucleotide polymorphism genotyping and point mutation detection by ligation on microarrays.
abstract: Based on the principle of sequencing by ligation, a novel method referred to as "Minisequencing by ligation" for detecting known single nucleotide variant in high-throughput assay formats is reported in this article. By designing a sequencing primer and fluorescently labeled probes each with a "discriminating" base according to the single base variation, this method can be used to analyze single nucleotide polymorphism (SNP) or point mutation in a number of samples in parallel. In our current study, three known nucleotides at a given position of three DNA templates with different CG contents are firstly genotyped on aldehyde-modified slide to testify the feasibility and optimize the reaction conditions. Then, by performing ligation reaction between phosphorylated 5' end of the sequencing primer and hydroxylated 3' end of the labeled probe or phosphorylated 5' end of the labeled probe and hydroxylated 3' end of the sequencing primer, a SNP locus (rs1800497 (C/T)) and a point mutation (Mt1555 (A/G)) have been accurately detected on aldehyde-modified microarray and Sepharose beads in acrylamide gel, respectively. It has demonstrated that "minisequencing by ligation," as a promising methodology, can perform point mutation and SNP analysis in a simple, cost-effective, robust and high-throughput way.
authors: State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, PR China.
authors: Yanqiang
authors: Y
authors: Li
authors: Jing
authors: J
authors: Tang
authors: Zhiqiang
authors: Z
authors: Pan
authors: Pengfeng
authors: P
authors: Xiao
authors: Dongrui
authors: D
authors: Zhou
authors: Lu
authors: L
authors: Jin
authors: Min
authors: M
authors: Pan
authors: Zuhong
authors: Z
authors: Lu
mesh_terms: D001483:Base Sequence; D011088:DNA Ligases; D004252:DNA Mutational Analysis; D017931:DNA Primers / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics; D005838:Genotype; D006801:Humans; D022521:Ligase Chain Reaction; D036103:Nanotechnology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA
chemical_list: D017931:DNA Primers; D004272:DNA, Mitochondrial; D011088:DNA Ligases