SNP基因分型的高通量方法

Chapter16

High-Throughput Methods for SNP Genotyping

Chunming Ding and Shengnan Jin

Abstract

Single nucleotide polymorphisms(SNPs)are ideal markers for identifying genes associated with complex diseases for two main reasons.Firstly,SNPs are densely located on the human genome at about one SNP per approximately500–1,000base pairs.Secondly,a large number of commercial platforms are available for semiautomated or fully automated SNP genotyping.These SNP genotyping platforms serve different purposes since they differ in SNP selection,reaction chemistry,signal detection,throughput,cost,and assay flexibility.This chapter aims to give an overview of some of these platforms by explaining the technologies behind each platform and identifying the best application scenarios for each platform through cross-comparison.The readers may delve into more technical details in the following chapters.

Key words:Whole genome association,fine mapping,single nucleotide polymorphism,copy number variation,haplotyping.

1.Introduction

Single nucleotide polymorphisms(SNPs)are best known as

genetic markers in disease-association studies to identify genes

associated with complex diseases(1,2).However,SNPs are also

used in many other clinically and biologically important applica-

tions(3).A large variety of commercial platforms are available for

semiautomated or fully automated SNP genotyping analysis.On

the basis of the purposes of the study,SNP genotyping can be

divided into two domains:whole genome association(WGA)and

fine mapping(Fig.16.1).Most of the genotyping platforms can

be classified accordingly.This chapter aims to briefly explain the

principles behind various platforms which lead to a comparison of

these platforms so that the readers will get a quick overview before

delving into the technical details of some of these methods in the

following chapters.

A.A.Komar(ed.),Single Nucleotide Polymorphisms,Methods in Molecular Biology578,

DOI10.1007/978-1-60327-411-1_16,ªHumana Press,a part of Springer Science+Business Media,LLC2003,2009

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2.Chemistries and Detection Methods for SNP Genotyping

Over the years,a number of chemistries were developed for dis-tinguishing two alleles of a SNP.The key for their adoption in high-throughput studies is dependent on the suitability for auto-mation.An ideal chemistry has to be universally applicable to any SNP (or to a substantial proportion of all human SNPs).Addi-tionally,high automation demands minimum steps in genotyping.

It may be fair to say that no single SNP genotyping platform is good enough to serve all purposes.

Generally,the chemistries for SNP genotyping can be roughly divided into two types based on the key reaction allowing for the SNP detection:(1)nonenzymatic differential hybridization (see Chapters 18and 19in this volume);(2)enzymatic reactions (see Chapter 23in this volume).Differential hybridization relies on different melting tempera-tures for matched and mismatched probes binding to the target DNA sequences.The Affymetrix SNP microarray employs this principle.For each SNP,four to six probes (25-mers each)are used.Affymetrix arrays can achieve very high density to accommodate millions of probes on a single chip.The newest Affymetrix Human SNP Array 6.0contains probes for 906,600SNPs and an additional 946,000probes for asses-sing copy number variations (CNVs).All the few million probes will be hybridized to their target sequences under the same temperature and buffer condition for the same amount of time,which is ideal for automated high-throughput SNP genotyping.However,the probes have to be effective in

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SNP Number

Sample Size

Fig.16.1.An overview of platforms with regard to throughput of single nucleotide polymorphisms and sample size.Platforms are selected on the basis of reasonable running costs.

246Ding and Jin