摘要
为研究SpHtp1基因在不同种属水霉菌株中的分布,以寄生水霉ATCC200013TM的基因组DNA为模板,设计1对特异性引物,进行PCR扩增。将扩增片段回收,并克隆入T载体,转化大肠杆菌DH5α,挑取阳性克隆测序。将测序结果与GenBank中登录的SpHtp1基因序列进行比较,同源性为99%。以该方法调查从水霉病主要流行地区分离的17株水霉菌SpHtp1基因的分布情况,结果发现:在17株水霉菌株中,有8株水霉菌检测到SpHtp1基因,阳性率为47%,且所有含SpHtp1基因的菌株在分类地位上均隶属于寄生水霉(Saprolegnia parasitica)。SpHtp1基因可能在寄生水霉中特异性存在,该基因有望用于寄生水霉的分离鉴定。
In order to understand the distribution of the SpHtp1 gene among different saprolegnia strains,using Saprolegnia parasitica(ATCC200013TM)genomic DNA as the template,apair of genespecific primers was designed,a PCR method was developed and the parameters for PCR amplification was optimized.The PCR product was extracted,linked into pMD18-T vector and then cloned into E.coli DH5α.The recombination plasmid was identified by PCR and sequenced to prove the amplification of the target gene.As a result,a 224bp DNA fragment of the SpHtp1 gene can be amplified from the saprolegniagenome,which had an identity of 99% in sequence with that of other Saprolegniastrains.Additionally,17 samples from all over China were tested using this method and the SpHtp1 gene was present in 8(47%)samples.The results indicated that the SpHtp1 gene is expressed specifically in S.parasitica,which can be used potentially for detection of Saprolegnia.
出处
《华中农业大学学报》
CAS
CSCD
北大核心
2014年第5期99-104,共6页
Journal of Huazhong Agricultural University
基金
国家"863"项目(2011AA10A216)
公益性行业(农业)科研专项(201203085)
上海市科技兴农重点攻关项目(沪农科攻字(2011)第4-8号)
国家自然科学基金项目(31172430)
国家水产种质资源平台运行服务项目
上海高校知识服务平台项目
