PCR-RFLP鉴别驽巴贝斯虫和马泰勒虫方法的建立

Establishment of PCR-RFLP Method for Identification of Babesia Caballi and Theileria Equi

马梨形虫病是一种由驽巴贝斯虫和马泰勒虫引起的蜱传血液寄生虫病,对我国养马业造成了巨大的经济损失。为了快速准确地鉴别驽巴贝斯虫和马泰勒虫,设计马梨形虫18S rRNA基因通用引物,运用PCR方法扩增序列,经过序列比对与分析,选用限制性内切酶DraⅠ对马梨形虫感染样本进行酶切。结果表明,马梨形虫两种病原18S rRNA基因PCR扩增产物长度为1500 bp。酶切后驽巴贝斯虫18S rRNA基因序列没有变化,马泰勒虫被切成大小为350 bp和1100 bp的两段,混合感染样本则形成350,1100 bp和1500 bp的3条带。实验建立的鉴别马梨形虫PCR-RFLP的方法实验流程简单,节约实验成本,可以快速并准确的鉴别马梨形虫的种类,为马梨形虫的分类提供了新的方法。

Equine piroplasmosis was a tick-borne blood parasitic disease caused by Babesia caballi and Theileria equi,which caused great economic losses to the horse breeding industry in China.In order to develop a taxonomic tool for equine piroplasm,the universal primers for 18S rRNA gene were designed,and the gene fragments were amplified for using PCR.After sequence alignment and analysis,the DraⅠwas chosen to identify the B.caballi and T.equi.The results showed that the length of PCR product of 18 S rRNA gene was 1500 bp.The enzyme digestion results of T.equi were digested into two segments of 350 bp and 1100 bp,B.caballi remain undigested,and the co-infection samples formed three bands of 350 bp,1100 bp and 1500 bp.The PCR-RFLP method established in this experiment for identification of equine piroplasm was simple in experimental process,saved experimental cost,and could quickly and accurately identify the species of equine piroplasm,which provided a new method for the classification of equine piroplasm.