旋毛虫肌幼虫RNA的提取及目的基因的PCR扩增

用改进的ＡＧＰＣ法成功地提取出旋毛虫肌幼虫总ＲＮＡ。每０．１ｍｌ压积虫体可获得１００μｇ的ＲＮＡ，其Ａ２６０与Ａ２８０及Ａ２６０与Ａ２３０的比值均接近于２，变性琼脂糖凝胶电泳显示，旋毛虫肌幼虫总ＲＮＡ缺少大部分真核生物所具有的２８Ｓ ｒＲＮＡ．通过比较研究，筛选出理想的虫体处理方法。用聚合酶链式反应直接从总ＲＮＡ中进行目的基因的扩增，得到一分子量为０．７ｋｄ的ＤＮＡ。通过限制性内切酶对其消化鉴定，

EXTRACTION OF RNA OF TBICHINELLA SPIRALIS MUSCLE LABVAE AND TARGET GENE AMPLIFICATION WITH PCR

The total RNA was extracted from T. spiralis (TSP) muscle larvae with an impovedAGPC technique. One hundred micrograin RNA couldbe obtained from 0.1ml of TSP.The A260/A280 and A260/A230 ratios of isolated RNA were approximately 2. Revealed bydenaturing agarose gel electrophoresis, the TSP total RNA was lack of 285 rRNA whichlocates in most of eukaryotic cells. The best method breaking TSP was screened bymaking a comparison among the three designs. The amplification of target gene wasdirectly Performed from total RNA using RNA PCR technique. The molecular weight ofgenerated cDNA was 0.7kb. Digesting it with the restriction enzymes, it was shown thatthe restriction sites of the obtained fragment were just as same as that of the knownsequence.