遗传多态性知识一、SNP, LD, Haplotype and Tagger SNP1. 遗传/基因多态性(genetic/gene polymorphism)在一随机婚配的群体中，染色体同一基因座位点上有两种或两种以上的基因型，且各个等位基因在群体中的出现频率皆高于1%。它是决定人体对疾病易感性、临床表现多样性及药物治疗反应差异性的重要因素。而种群中频率等于或小于1 %的碱基变异称为突变。染色体同一DNA位置上的每个碱基类型叫做一个等位位点。如某些人的染色体上某一位置的碱基是A，而另一些人的染色体上相同位置上的碱基是G，除性染色体外,每个人体内的染色体都有两份,所以,一个人所拥有的一对等位位点的类型被称作基因型(genotype)，如GA、GG、AA；检定一个人的基因型，被称作基因分型(genotyping)。由不同基因型与环境共同作用所产生的生物体（人类）可观测的物理或生理性状称为表现型（phenotype）。限制性片段长度多态性(restriction fragment length polymorphism. RFLP)是第一代的遗传标记；可变数目的串联重复(variable number of tandem repeat. VNTR)是第二代遗传标记；其中重复单位为2-6个核苷酸称为微卫星或短串联重复；6-12个核苷酸称为小卫星。Polymorphisms are defined as frequent (occurring in greater than 1% of the population) variations in the human DNA sequence. Most involve a single base pair substitution, known as single nucleotide polymorphisms(1), although more complex variations are also recognised. SNPs are single base pair positions in genomic DNA at which different sequence alternatives (alleles) exist in normal individuals in some population(s), wherein the least frequent allele has an abundance of 1% or greater. In principle, SNPs could be bi-, tri-, or tetra-alletic polymorphisms. Howere, in humans, tri-alletic and tetra-alletic SNPs are rare almost to the point of non-existence, and so SNPs are sometimes simply referred to as bi-allelic markers. 单核苷酸多态性(single nucleotide polymorphism.SNP)：最早由美国麻省理工学院的人类基因组研究中心Lander于1996年提出，是不同个体基因组DNA序列内特定核苷酸位置上单个碱基的不同是第三代遗传标记，任一SNP在群体中出现的频率应不小于1%，原则上SNP可以是双、三、四等位基因多态，在人类三、四等位基因的SNP很少甚至几乎不存在，因此SNP简单指双等位标记，双等位基因的SNP替换包括1个转换CT(GA)和3个颠换CA(GT)、CG(GC)、TA(A T)，由于核苷酸的5-甲基胞嘧啶脱氢基反应相对比较频繁，使得四种SNPs在基因组中出现的频率不同，在生物体内约2/3是C/T(G/A)转换，并且多存在于非转录序列中。据统计，人类基因组中3*109碱基中至少存在着1000万个SNPs位点，平均约1个SNP/1000bp。与其他遗传标记(如限制性片段长度多态，短串联重复)的主要不同是不再以“长度”的差异作为检测的手段，而直接以序列的变异作为标记，具有高丰度、高度稳定性和易于自动化分析等独特的优势。英文描述：SNP markers are preferred over microsatellite markers for association studies, because of their high abundance along the human genome (SNPs with minor allele frequency0.1 occur once every 600 kb) (Wang et al.1998), their low mutation rate, and the accessibility of high-throughput genotyping. The power of association studies based on SNPs depends not only on the sample size and density of the marker map but also on many other factors, such as the age and frequency of the disease mutations and SNPs and the extent of linkage disequilibrium(LD) in the region.(2)根据SNP在基因序列中所处的位置的不同，SNP位点可以分为几个大类。大多数对基因的功能没有影响的SNPs,称为anonymous SNPs；存在于基因内部的SNP位点则称为gene-based SNPs，包括内含子、外显子和启动子中的单核苷酸多态性位点。其中，存在于蛋白质编码序列中的SNP位点称为cSNPs或coding SNPs。在cSNPs中，如果不改变所编码的氨基酸序列，这样的单核苷酸多态性称为synonymous SNPs；如果SNP导致了氨基酸序列的改变，则称为non-synonymous SNPs。发生在基因蛋白编码区的SNP，可能引起编码氨基酸的置换，导致蛋白功能的改变；大多数SNPs发生在非编码区，启动子区域的SNP也许影响转录因子结合的能力，改变基因转录的速率或水平；发生在5上游区或3下游区域的SNPs可能改变转录的mRNA的稳定性或增强子活性；而内含子区域的SNPs的功能效应有待于进一步研究(3)。检测SNP的方法多种多样，有直接测序法、PCR-RFLP法、单链构型多态分析法(single strand conformation polymorphism analysis，SSCP)、异源双链分析法(heteroduplex analysis，HA)、变性梯度凝胶电泳分析法(denaturing gradient gel electrophoresis，DGGE)、固相化学断裂法(solid phase chemical cleavage method，spCCM)、等位基因特异性聚合酶链反应法(allele-specific PCR)、DNA芯片检测法和实时荧光定量PCR法等，均具有较高的特异性和敏感性，不同实验室可以根据研究目的和经费选择合适的检测方法。2. 单倍型(haplotype)位于染色体上特定区域、相互关联、倾向于以整体模式遗传给后代的SNPs组合称作单倍型(haplotype)，比拟为人类进化历史的“分子化石”。在一段DNA内若存在n个SNP位点，则群体内理论上可能存在2n种单倍型，但针对每一个体来说只有2种单倍型。单倍型构建方法：实验方法目前有单分子稀释法(single-specific dilution)、AP-PCR(allele-specific PCR)、长插入克隆法(Long-insert cloning)与双倍型-单体型转化(diploid-to-haploid conversion)等；统计算法有Clark算法、最大似然算法、贝叶斯算法。3. 单倍域(haplotype block)根据基因组大范围内SNPs之间的连锁不平衡，能够用一个相对简单的模型来描述人类基因组的单倍型结构，即染色体上存在的连续的、稳定的、几乎没有被重组所打断的单倍型区域，称为单倍域(haplotype block or haploblocks)。Several neighboring, tightly linked SNPs are inherited together and form a haplotype block.单倍域可能是遗传的最小单位，在极端情况下，它可以是一个单独的SNP或者是一整条染色体，重组事件频发的区域可将相邻的单倍域间隔开来。3.1 单倍域的定义：a haplotype block is a contiguous set of markers in which the average D(the standardized coefficient of LD(4) is greater than some predetermined threshold. Gabriel et al(5) described human genome can be parsed objectively into haplotype blocks: sizable regions over which there is little evidence for historical recombination and within which only a few common haplotypes are observed. based on linkage disequilibrium (LD), that is large pairwise |D| values between those SNP pairs within one haploblock. Patil et al(6) defined haplotype blocks as a region with a large proportion(80%) of inferred common haplotypes.based on the concept of “chromosome coverage” , with a haplotype block containing a minimum number of SNPs that account for a majority of common haplotypes or a reduced level of haplotype diversity.Wang et al(7) further proposed explicit“no historical recombination” as a definition for haplotype blocks, which can be tested using a four-gamete test.Ding K et al(8) choose to define haplotype blocks based on LD when haplotype-block-based tSNPs selection methods were employed. The LD-based haplotype-block definition requires that the proportion of SNP pairs with strong D(absolute D0.70) must account for