干扰日本血吸虫凋亡抑制因子最佳siRNA分子的筛选  

Screening of the siRNAs for silencing inhibitor apoptosis protein in Schistosoma japonicum

作　　者：罗荣[1] 赵江平[1] 胡超[1] 程国锋[1] 

机构地区：[1]中国农业科学院上海兽医研究所,农业部动物寄生虫学重点开放实验室,上海200241

出　　处：《中国人兽共患病学报》2013年第9期846-849,共4页Chinese Journal of Zoonoses

基　　金：国家自然科学基金(No.30901068)资助~~

摘　　要：目的筛选干扰日本血吸虫凋亡抑制因子(Inhibitor of apoptosis protein of Schistosoma japonicum,SjIAP)较好的siRNA分子。方法应用生物信息学设计了针对SjIAP的3对siRNA分子。利用脂质体将每对siRNA分子和表达IAP的重组质粒传染到HEK293T细胞中,利用实时定量RT-PCR和免疫印迹检测IAP的表达。将siRNA分子与日本血吸虫童虫(12d)进行体外共培养,利用实时定量RT-PCR和免疫印迹检测虫体内的IAP表达。结果细胞转染实验的实时定量RTPCR和免疫印迹分析表明本研究获得了2个干扰SjIAP较好的siRNA分子,且其中1对siRNA分子可显著抑制日本血吸虫虫体内IAP蛋白的表达,同时虫体Caspase活性有所上升。结论本研究获得了2个干扰日本血吸虫IAP较好的siRNA分子,为进一步利用RNA干扰研究血吸虫IAP的功能奠定了前期基础。Our previous study suggests that the inhibitor apoptosis protein in Schistosoma japonicum （SjlAP） may play important roles in parasitic living and development as well as in the host parasite interactions. To obtain the best efficacy of small interfering RNA （siRNA） for silencing the SjIAP, three siRNA duplexes targeting SjlAP were bioinformatically de signed and chemical synthesized. Then, each siRNA duplex and the recombinant plasmid for expressing SjIAP were co-trans fected into human HEK293T cells. Real-time RT-PCR and Western blot analyses indicated that the siRNA 951 showed the best efficacy for silencing SjlAP expression in both mRNA and protein levels. Then, the siRNA 95l was further added into the me dium of in vitro cultured schistosomula （12 days）. Western blot analysis indicated that the expression of SjlAP to some extent inhibited at the protein level. In addition, the activities of Caspase 3/7 were also increased in the worms treated with siRNA 951. In conclusion, the best efficacy of siRNA for silencing SjlAP was obtained and the present study provides foundational in- formation for the further investigation on SjIAP functions by using RNA interference.

关 键 词：日本血吸虫 凋亡抑制因子 RNA干扰 基因功能 

分 类 号：R383.2[医药卫生—医学寄生虫学]