半滑舌鳎性别特异微卫星标记的SCAR转化及其应用  

SCAR-transformation of Sex-specific SSR Marker and Its Application in Half-smooth Tongue Sole(Cynoglossussemiliaevis)

作　　者：刘洋[1,2] 陈松林[1] 高峰涛[1] 孟亮[1] 胡乔木[1] 宋文涛[1] 邵长伟[1] 吕为群[2] 

机构地区：[1]中国水产科学研究院黄海水产研究所、农业部海洋渔业可持续发展重点实验室,青岛266071 [2]上海海洋大学水产与生命学院,上海201306

出　　处：《农业生物技术学报》2014年第6期787-792,共6页Journal of Agricultural Biotechnology

基　　金：国家高技术研究发展计划(863)项目(No.2012AA10A403-2);国家自然科学基金项目(No.3113005和No.41006107);山东省泰山学者工程专项

摘　　要：半滑舌鳎(Cynoglossus semiliaevis)性别控制与全雌育种相关研究需要一种快速、准确的性别特异分子标记。本研究通过对半滑舌鳎共显性性别特异微卫星标记筛选,得到Z、W染色体同源片段相差35bp的位点,对其进行克隆和测序,针对所得测序结果,重新设计引物进行序列特异扩增区间(sequence characterized amplified region,SCAR)转化,增大35 bp差异片段在扩增片段中的所占比例;并摸索得到4%琼脂糖凝胶,150 V,25 min为最适电泳条件,所得SCAR标记可在雌性个体中扩增出169和134 bp两种DNA条带,雄性个体中扩增出169 bp DNA条带,超雌个体中扩增出134 bp DNA条带;将其应用于生产实践中,对半滑舌鳎生理雄性亲鱼576尾进行快速检测,在保证存活的前提下,剔除了亲鱼中的100尾伪雄鱼,有助于提高后代的雌性比例。本研究所得半滑舌鳎性别特异SCAR标记,可用于实验室对半滑舌鳎雌性、雄性和超雌个体遗传性别的准确测定和养殖场简易环境下养殖亲鱼雄鱼中伪雄个体的快速检测。A fast and exact sex-specific molecular marker is important in the study of sex control and allfemale breeding in half-smooth tongue sole（Cynoglossus semiliaevis）. In the present study, a sex-specific codominace simple sequence repeat（SSR） marker which had 35 bp difference fragment between the isogeny area of Z and W chromosone was filtrated from 37 same type markers. The selected marker was cloned andsequenced, then the sequence characterized amplified region（SCAR） primer was designed to increase the proportion of the 35 bp defference fragment. Interrelated analyses concluded that 4% agarose gel, 150 Velectrophoresis for 25 min were the optimum condition. The transformed SCAR marker could amplificate 134, 134 and 169, 169 bp fragment in super-female, female and male individual, respectively. The SCARmarker was applied in production practice, and 574 physiological male parent half-smooth tongue sole were tested and 100 neo-males were identified and got rid of in less harmful condition which was helpful toincrease the female proportion of the offspring. The SCAR marker can be used in accurate test of female, male and super-female individual in laboratory and fast test of male parent half-smooth tongue sole to get rid of neo-male individual under simple and crude environment like fish farm.

关 键 词：半滑舌鳎 性别特异标记 序列特异扩增区间(SCAR) 快速检测 

分 类 号：S512.1[农业科学—作物学]