褐牙鲆耐热相关分子标记筛选及遗传多样性分析

The screening of molecular markers correlated to thermal-tolerance and genetic diversity analysis of selective stocks of Japanese flounder(Paralichthys olivaceus)

通过高温刺激(32℃)获得褐牙鲆的耐热群体和热敏感群体,利用扩增片段长度多态性(amplified fragment length polymorphism,AFLP)和微卫星技术筛选与褐牙鲆耐热性相关的分子标记,并分析了两个群体的遗传多样性。采用81对AFLP引物组合和42对微卫星引物进行耐热相关分子标记的筛选,结果表明:1个AFLP位点(A3)在热敏感群体中出现的频率显著高于耐热群体,与耐热性存在显著的负相关(P<0.01),相关系数达到-0.453;4个AFLP位点在耐热群体中出现的频率显著高于热敏感群体,与耐热性存在显著正相关性(P<0.01);微卫星引物Po25A和205TUF扩增出的位点S1和S2与耐热性存在显著负相关性(P<0.01),相关系数分别为-0.408和-0.398。群体遗传多样性分析表明耐热群体与热敏感群体的遗传变异水平较高,两个群体的遗传多样性水平相当。研究的结果不仅为褐牙鲆的耐高温选育提供技术支持,而且可以为鱼类抗逆性研究提供参考数据。研究亮点:利用AFLP和微卫星技术筛选褐牙鲆耐热相关的分子标记并分析其群体的遗传多样性。研究的结果不仅为褐牙鲆的耐高温品系选育提供技术支持,而且可以为鱼类抗逆性研究提供参考。

The studied Japanese flounders （Paralichthys olivaceus） were divided into two categories： thermaltolerance stock and thermal-susceptible stock by exposing in heated water （32℃ ）. Amplified fragment length polymorphism （AFLP） and microsatellite technology were employed to screen the molecular markers correlated to thermal-tolerance and analyze the genetic diversity of thermal-tolerance stock and thermalsusceptible stock of Japanese flounder. Eighty-one AFLP primer combinations and forty-two microsatellite primers were used to screen molecular markers correlated to thermal tolerance of Japanese flounder. AFLP and microsatellite loci which occurred in different genotype frequencies were analyzed using XE-test to examine the correlation between the molecular markers and thermal tolerance in Japanese flounder. The results showed that the frequency of AFLP locus A3 was significantly higher in thermal-susceptible stock than that in thermaltolerance stock, which indicated that this locus was negatively correlated with thermal tolerance, and the parameter of correlation was - 0. 453 （ P 〈 0.01 ）. Also, the frequencies of 4 AFLP loci were signicantly higher in thermal-tolerance stock than those in thermal-susceptible stock, which indicated that the 4 loci were correlated with thermal tolerance （P 〈 0.01 ）. Based on microsatellite analysis, the loci S1 and S2 amplified by Po25A and 205TUF showed to be negatively correlated with thermal tolerance, and the parameters of correlations were -0. 408 （P 〈 0. 01） and -0. 398 （P 〈 0. 01 ）, respectively. In addition, the genetic diversity of thermal-tolerance stock and thermal-susceptible stock were analyzed using eight AFLP primer combinations and six microsatellite primers. The results showed high genetic variations in the two stocks, and the genetic diversity of the two stocks was comparable. The results of present study could be helpful for the selective breeding of heat-resistant flounders and provide reference basis for the stress resistance study of fish.