小叶杨DREB同源基因内SSR的发现及群体结构分析

利用基因特异的PCR引物从小叶杨(Populus simonii)经干旱胁迫后构建的cDNA文库中扩增得到长度为671bp的PsDREB cDNA克隆。该基因内部含有完整的且大小为543bp的开放阅读框,可编码181个氨基酸残基。在此基础上,对36株小叶杨基因型个体的DREB基因进行了克隆和序列分析,发现其内部存在以CAC为基本单位的4次(A)、5次(B)、7次(C)、8次(D)、9次(E)、10次(F)和11次(G)7种类型的简单重复序列(SSR)。以该重复序列为扩增对象设计引物,对我国11个省16个群体528株小叶杨进行了群体遗传结构分析。结果表明,在检测群体中各位点等位基因频率大小顺序依次为:DCFEAGB;平均有效等位基因数为3.1670,平均观察和期望杂合度分别为0.4685和0.5315,且在多数群体中存在一定程度的近交效应和显著偏离Hardy-Weinberg平衡;而Ewens-Watterson中立性检验表明,除山西宁武、辽宁朝阳和河南伊川外,其它多数群体均属于中立性选择。研究结果将为利用候选基因内SSR标记来评价林木育种资源的遗传多样性以及保护和利用这些资源提供科学的理论依据。

DREB Gene and Its Application in Analyzing Population Structure in Populus simonii

A full-length cDNA clone encoding DREB was isolated from a cDNA library prepared under drought stress by the gene-specific primer amplification from Populus simonii. The cDNA was 671 bp with an open reading frame (543 bp) capable of encoding a protein of 181 amino acids. The DREB genomic sequences of 36 unrelated individuals were cloned and analyzed. A trinucleotide repeat motif of the CAC structural unit was detected within the coding region of DREB. This simple sequence repeat (SSR) was of 7 types, including 4(A), 5(B), 7(C), 8(D), 9(E), 10(F) and 11(G) times. To evaluate the diversity of the target SSR in the natural populations of P. simonii, SSR specific primer pairs were used to test the genetic structure in 16 natural populations of 528 individuals covering 11 provinces of China. The frequency distribution of the SSR allele from high to low was D＞C＞F＞E＞A＞G＞B in 16 populations. With mean effective number of alleles of 3.167 0, the mean value of observed heterozygosis and expected heterozygosis of the SSR allele was 0.468 5 and 0.531 5, respectively. Most populations with this SSR loci exhibited more or less inbreeding effects and deviated from Hardy-Weinberg equilibrium. Except for the populations of Ningwu (Shanxi province), Chaoyang (Liaoning province) and Yichuan (Henan province), this locus was neutral in all the other populations by Ewens-Watterson central testing. These results provide an important theoretical foundation for evaluating genetic diversity of germplasm resources with SSR markers within candidate genes and its application in conservation and use of breeding resources in forest trees.