摘要
细粒棘球绦虫原头蚴(PSC)具有双向发育的特点,本文旨在利用SSH和生物信息学方法筛选原头蚴包囊(CW)和成虫(AW)特异性的基因。将PSC-CW和PSC-AW双相发育差减cDNA文库的差减PCR产物克隆入pGEM-T载体并测序,利用CAP3sequence assembly在线软件、Blast2GO软件与GenBank数据库对测序结果进行分析,筛选原头蚴成囊和成虫特异性的基因,并探讨这些基因的功能。结果显示:对PSC-CW和PSC-AW文库进行扩增,分别得到280和200个阳性克隆,菌落PCR鉴定结果表明,这些克隆均插入200~1 000bp片段。测序结果显示从PSC-CW和PSC-AW SSH文库中分别得到16和10个特异性基因。其中PSC-CW SSH文库中得到4个出现频率较高的基因,其余12个基因仅出现1次,其中5个为未知基因,已知基因分别编码细胞色素C氧化酶Ⅰ、热休克蛋白70和铁蛋白等功能蛋白质。PSC-AW SSH文库中得到10个基因,其中2个出现频率较高基因,8个基因仅出现1次,4个为未知基因,6个已知基因分别编码基质蛋白1、延伸因子1α和脂肪酸结合蛋白等功能蛋白质。本研究筛选获得原头蚴成囊和成虫发育时差异表达的基因,对这些基因的功能进行分析表明,PSC-CW SSH文库中多是与营养和能量转运功能的相关基因,而PSC-AW SSH文库中多为发育与分化相关的基因,这些基因的差异表达可能与原头蚴处于不同的生长环境和发育方向有关,同时也为棘球蚴病免疫诊断、药物筛选和疫苗研制提供候选基因。
According to the bidirectional development of the protoscolex(PSC)of Echinococcus granulosus,the study was conducted to screen the specifical genes of E.granulosus PSC development between a larval cyst and adult worm.The subtractive PCR products of the SSH library ofE.granulosus PSC-adult(AW)and E.granulosus PSC-cyst(CW)were connected to the pGEMT easy vector,and were cloned and sequenced.Totally,280 and 200clones were obtained from PSC-CW and PSC-AM libraries,respectively,with the length of fragments 200 to 1000bp.The clones were sequenced.Sequence alignment analysis showed that these sequences belong to 16and10 gene clusters representing 16 and 10unique genes from PSC-CM and PSC-AW libraries respectively.As the genes are specifically or differentially expressed during the development of PSC to either adult worm or cyst,they are important for the parasite development and differentiation,thus can be candidates for drug and vaccine development.
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2015年第7期1215-1223,共9页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
公益性行业(农业)科研专项项目(201103008)
关键词
细粒棘球绦虫
抑制差减杂交
特异性基因
Echinococcus granulosus
suppression subtractive hybridization(SSH)
the specifical genes
