摘要
以尼罗罗非鱼(Oreochromis niloticus)为对象,采用体内注射植物血细胞凝集素(PHA)方式收集有丝分裂后期的鱼体头肾细胞,低渗法制片,运用激光显微分离获得尼罗罗非鱼1号染色体,分离后的单染色体通过特异性酶切和LA-PCR体外扩增,得到500~2 000 bp之间的DNA片段,并通过Southern blot和微卫星标记得到验证。扩增片段通过电泳回收、连接转化、感受态细胞培养等步骤构建尼罗罗非鱼1号染色体微克隆文库,克隆片段长度在300~600 bp之间。
The head-kidney cells of Oreochromis niloticus were collected by phytohemagglutinin （ PHA） injection in vivo and the metaphase chromosome specimens were prepared by the low osmotic method.The chromosome 1 of O.niloticus was successfully separated by laser microdissection method.Meanwhile,the generated DNA fragments ranging from 500 to 2 000 bp were amplified by specific enzyme cutting and fleetly amplification of linker adaptor mediated PCR （ LA-PCR）.The amplified products of single chromosome were verified by Southern blot and microsatellite markers.SSRs in the amplified fragments were detected with developed specific SSR primers.A genome clone of O.niloticus chromosome 1 was constructed by electrophoresis recycling,transformation and cell cultivation.The constructed micro cloning library of the chromosome 1 in O.niloticus contains fragments range from 300 to 600 bp.
作者
耿青
肖炜
李大宇
邹芝英
祝璟琳
杨弘
GENG Qing;XIAO Wei;LI Da-yu;ZOU Zhi-ying;ZHU Jing-ling;YANG Hong(Wuxi Fisheries College,Nanjing Agricultural University,Wuxi 214081,Jiaugsu,China;Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization,Ministry of Agriculture;Freshwater Fisheries Research Center,Chinese Academy of Fishery Sciences,Wuxi 214081,Jiaugsu,Chin)
出处
《淡水渔业》
CSCD
北大核心
2018年第4期3-8,共6页
Freshwater Fisheries
基金
现代农业产业技术体系专项资金资助(CARS-46)
