SNP discovery by amplicon sequencing and multiplex SNP genotyping in the allopolyploid species Brassica napus
SNP discovery by amplicon sequencing and multiplex SNP genotyping in the allopolyploid species Brassica napus
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DOI：
10.1139/g10-079
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发表时间：
2010-11-01
期刊：
影响因子：
3.1
通讯作者：
Ganal, M. W.
中科院分区：
文献类型：
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作者：
Durstewitz, G.;Polley, A.;Ganal, M. W.
油菜（Brassica napus）是由两个基因组组成的异源四倍体物种，分别来源于B. rapa （A基因组）和B. oleracea （C基因组）。这两个基因组的存在使得单核苷酸多态性（SNP）标记的鉴定和分析比二倍体物种更具挑战性，因为对于给定的位点，在SNP鉴定和分析过程中通常必须同时分析两个版本的DNA序列（基于两个祖先基因组）。研究人员分析了来自表达序列标签（est）的100个扩增子，以鉴定一组油菜品种和代表祖先基因组的两个姊妹种中的snp。共鉴定出604个SNP，平均每42 bp有1个SNP。可以清楚地区分不同植物品种之间的多态性snp和区分两个祖先基因组的snp。为了验证已鉴定的snp在遗传分析中的应用，我们开发了Illumina GoldenGate检测一些已鉴定的snp。通过对不同油菜品种的分析，并利用GoldenGate技术对油菜群体进行定位，我们能够在异源四倍体油菜中识别出许多不同的分离模式。大多数鉴定的SNP标记可以很容易地用于遗传定位，这表明扩增子测序和Illumina GoldenGate检测可以可靠地鉴定四倍体油菜的SNP标记，并将其转化为可用于遗传分析的成功SNP检测。
Oilseed rape (Brassica napus) is an allotetraploid species consisting of two genomes, derived from B. rapa (A genome) and B. oleracea (C genome). The presence of these two genomes makes single nucleotide polymorphism (SNP) marker identification and SNP analysis more challenging than in diploid species, as for a given locus usually two versions of a DNA sequence (based on the two ancestral genomes) have to be analyzed simultaneously during SNP identification and analysis. One hundred amplicons derived from expressed sequence tag (ESTs) were analyzed to identify SNPs in a panel of oilseed rape varieties and within two sister species representing the ancestral genomes. A total of 604 SNPs were identified, averaging one SNP in every 42 bp. It was possible to clearly discriminate SNPs that are polymorphic between different plant varieties from SNPs differentiating the two ancestral genomes. To validate the identified SNPs for their use in genetic analysis, we have developed Illumina GoldenGate assays for some of the identified SNPs. Through the analysis of a number of oilseed rape varieties and mapping populations with GoldenGate assays, we were able to identify a number of different segregation patterns in allotetraploid oilseed rape. The majority of the identified SNP markers can be readily used for genetic mapping, showing that amplicon sequencing and Illumina GoldenGate assays can be used to reliably identify SNP markers in tetraploid oilseed rape and to convert them into successful SNP assays that can be used for genetic analysis.