小熊猫蛔虫ITS及5.8S rDNA序列的克隆与分析

以广州动物园小熊猫体内分离出的蛔虫为研究对象,运用PCR方法,以保守引物NC5和NC2扩增其核糖体DNA（rDNA）的内转录间隔区（ITS）和5.8S序列,并对扩增后的片段进行纯化、克隆至pGEM-Teasy载体、转化、测序和序列分析,以鉴定小熊猫蛔虫的种类。结果显示2条蛔虫样品的ITS及5.8S rDNA序列基本一致,总长为913 bp,样品间序列相似性为99.7%。将序列与GenBank^TM公布的相关序列进行比较分析,结果显示2条蛔虫的ITS及5.8S序列与黑熊横走贝蛔虫（Baylisascaris transfuga注册号AB571304）相似性分别为98.1%、98.4%,与大熊猫西氏贝蛔虫（Baylisascaris schroederi注册号JN210912）相似性分别为96.9%、97.1%,与猪蛔虫（Ascaris suum注册号AB571302）相似性分别为89.9%、90.1%,与人蛔虫（Ascaris lumbricoides注册号AB571296）相似性分别为89.8%、90.1%,ITS-1序列与浣熊贝蛔虫（Baylisascaris procyonis注册号AB053230）相似性分别为92.0%、92.3%。研究结果表明小熊猫体内分离的蛔虫可能为贝蛔属蛔虫,从而为蛔虫的进一步分类、鉴定和遗传变异研究奠定了基础。

Cloning and Sequence Analysis of ITS and 5.8S rDNA of Ascarid Nematodes from Red Panda(Ailurus fulgens)

Our research objective was to identify ascarid samples isolated from a red panda at the Guangzhou Zoo.The internal transcribed spacer(ITS) and 5.8S of ribosomal DNA(rDNA) of two ascarid samples were amplified by PCR using a pair of conserved primers(NC5 and NC2),and the amplified products were purified,cloned and sequenced.The lengths of sequences (ITS and 5.8S rDNA) from two samples were both 913 bp,and the similarity of sequences was 99.7%between the two ascarid samples.The similarities of ITS and 5.8S sequences were 98.1%and 98.4%,respectively,compared with the relevant sequences of Baylisascaris transfuga(AB571304) available from GenBank.Similarities were 96.9%and 97.1% compared with Baylisascaris schroederi(JN210912),89.9%and 90.1%compared with Ascaris suum(AB571302),89.8% and 90.1%compared with Ascaris lumbricoides(AB571296),and 92.0%and 92.3%compared with the ITS - 1 sequence of Baylisascaris procyonis(AB053230).We conclude that the ascarid nematodes might represent Baylisascaris sp.This research, provided a foundation for further studies of molecular identification and molecular genetics of ascarid.