摘要
针对编码猪蛔虫雌虫卵巢蛋白基因的EST序列设计了1对特异引物及连接T7启动子的引物，经PCR扩增，获得5’端连有T7启动子的双链DNA，在RNA聚合酶的作用下，转录合成dsRNA。通过浸泡的方式将dsRNA导入秀丽新杆线虫中，在浸泡后的5个不同的时间点，采用RT—PCR检测虫体浸泡后靶基因被干扰的效果。试验结果表明，在浸泡后的8～29h能检测到同源靶标的存在；而在浸泡后的43～57h不能检测到靶标RNA。另外，试验组没有观测到虫体明显的表型变化，而对照组在浸泡28h后，发现部分线虫体内有发育成形的虫卵。初步认为导入虫体的dsRNA发生了干扰效应。
Based on the sequence of EST F632 encoding A. suum female ovary protein, a pair of specific primers was designed to amplify the F632, and the T7 promoter sequence was added to 5＇ end of the primers, and the transcription template containing T7 sequence was generated. Transcription reaction was performed, and dsRNA was synthesized. The dsRNA was delivered into C. elegans by soaking, and RT- PCR was used to detect the target mRNA after different lengths of soaking time. In results, PCR products were amplified from the worms soaked for 8-29 hours with the dsRNA, but no PCR products were amplified after soaking for 43-57 hours, indicating that the internal mRNA was destructed. Phenotypical observation of the worm development revealed no significant changes in the experimental worms, whereas the egg development was observed for the control C. elegans after soaking for 28 hours. The results indicated that RNAi effect was evident.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2006年第9期724-728,共5页
Chinese Veterinary Science
基金
国家杰出青年科学基金项目(30225033)
广东省自然科学基金重点项目(36835)
广东省科技计划项目(2004B50201020)
关键词
猪蛔虫
性别基因
RNA干扰
功能
Ascaris suum
gender-specific genes
RNA interference
function
作者简介
邹丰才（1976-），男，四川成都人，博士，E-mail：zfc1207@163．com。
朱兴全为通讯作者，Tel：020-85283730，E-mail：xingquanzh@scau．edu．cn
