魁蚶4个地理群体ITS序列变异及系统发生分析

ITS sequence variation and phylogenetic analysis on four geographical populations of blood clam Scapharca broughtonii

为充分了解我国魁蚶种质资源状况,准确定位魁蚶的养殖育种模式,本研究采用聚合酶链式反应(PCR)对采自山东蓬莱(PL)、山东黄岛(HD)、江苏前三岛(QSD)及韩国统营(KTY)4个地理群体魁蚶样本的核糖体RNA两个内转录间隔区域(ITS-1和ITS-2)进行扩增,经测序比对后分别获得长度为468bp和541bp(包含引物、插入/缺失位点)的序列。序列比对结果表明,ITS区序列变异相对较高,但4个群体的碱基组成基本一致;4个群体的ITS区序列均表现出丰富的遗传多样性,HD群体最为丰富,该群体在ITS区序列上的变异位点数也最多,HD和QSD群体共同变异位点多且集中。对不同个体间的遗传距离和分子系统树分析结果表明,ITS序列在魁蚶不同地理群体间甚至个体间存在差异,且ITS-1和ITS-2的变异频率是不一样的,4个群体合计55个个体基于ITS-1序列没有发生明显聚类现象,这4个群体合计56个个体基于ITS-2序列明显聚为PL/KTY、HD/QSD两大类,基于ITS-2序列进行系统发生分析的结果同于课题组之前基于形态度量学分析和分子标记分析的结果;HD群体遗传多样性丰富,而KTY群体遗传变异相对较少,各群体之间存在一定的遗传差异,综合考虑不同群体相对优良的生物学特征,可开展选择或杂交培育良种的研究。

In order to investigate the genetic structure and variability of Scapharca broughtonii, thus to make proposals for the species culture and breeding, four geographical pop ulations of S. broughtonii were collected from Shandong Penglai（PL）, Shandong Huangdao HD）, Jiangsu Qiansandao （QSD） in China, and Tongying in Korea （KTY） respectively, and genetic variation was analyzed based on ITS nucleotide sequences of rDNA. PCR amplification products were sequenced and aligned subsequently. The 486bp and 541bp necleotide sequences of ITSq and ITS-2 respectively were obtained（primers and insertion/deletion sites included）. The results showed that though the genetic variation of ITS-1 and ITS-2 was relatively high, the base composition of them was stable. The genetic diversity of the four populations was rich, which indicated that the germplasm resource of the species was in good condition. The genetic diversity of HD population was the richest. Also, the number of variation sites of ITS-1 and ITS-2 were the largest in HD population. There were many neighbor common variation sites in HD and QSD populations. In this study, we calculated the genetic distances between individuals and constructed the molecular phylogenetic tree by using minimum evolution method by MEGA 4.0. The results showed that the nucleotides in one species＇ genome had different variation fre- quency,and variation of nucleotides segment couldn＇t represent the genetic variation of the whole genome. Fifty-five blood clams did not cluster obviously based on the ITS-1 segment base se- quences. However, 57 blood clams can be clustered into two catigories completely, one as HD and QSD population, the other as KTY and PI. populations. The information from nucleotides variation of ITS-2 is similar to the conclusion from multivariate morphomertrics and molecular marker analysis carried out by our research group in the past. The genetic diversity of HD population is very rich, while the KTY population showed relatively low variation. There were certain genetic differences between each population, so that cross- and selective-breeding may be tested according to their biological characteristics.