Rappelli P, Rocchigiani A M, Erre G, Colombo M M, Cappuccinelli P, Fiori P
Istituto di Microbiologia e Virologia, Università di Sassari, Italy.
FEMS Microbiol Lett. 1995 Jun 1;129(1):21-6. doi: 10.1016/0378-1097(95)00124-N.
A Trichomonas vaginalis cDNA library was constructed and recombinant plaques were screened using rabbit immunoglobulins specific for P65, a protozoan protein involved in pathogenicity that we identified in a previous study. A 1.38 kilobases cDNA fragment coding for the P65 protein was cloned in E. coli and then sequenced. On the basis of of the sequence obtained, six primers were synthesised and used to set up a Polymerase Chain Reaction. The presence of a specific amplicon in all 30 clinical isolates tested shows that P65 is a conserved and stable gene. The reaction is highly sensitive (as few as 5 to 10 parasites can be detected) and specific for Trichomonas vaginalis; the gene coding P65 adhesin can be therefore considered a very good molecular target for polymerase chain reaction-based diagnostic purposes.
构建了阴道毛滴虫cDNA文库,并用针对P65的兔免疫球蛋白筛选重组噬菌斑,P65是我们在先前研究中鉴定出的一种参与致病性的原生动物蛋白。编码P65蛋白的1.38千碱基cDNA片段克隆到大肠杆菌中,然后进行测序。根据获得的序列,合成了6条引物并用于建立聚合酶链反应。在所有检测的30个临床分离株中均存在特异性扩增子,表明P65是一个保守且稳定的基因。该反应高度敏感(可检测低至5至10个寄生虫)且对阴道毛滴虫具有特异性;因此,编码P65黏附素的基因可被认为是基于聚合酶链反应诊断目的的非常好的分子靶标。