Paces J, Urbánková V, Urbánek P
Institute of Molecular Genetics, Czechoslovak Academy of Sciences, Prague.
Mol Biochem Parasitol. 1992 Sep;54(2):247-55. doi: 10.1016/0166-6851(92)90116-2.
A family of 650-bp-long repeats from the Trichomonas vaginalis genome, designated the Tv-E650 family, was cloned and sequenced. The nucleotide sequence is A+T-rich (73.3% A+T in the consensus sequence) and highly conserved among the 8 molecular clones analyzed. The differences among the clones are single-nucleotide and 2-nucleotide substitutions and insertions or deletions. The sequence uniformity of the clones as well as the presence of identical mutations in different clones suggest that efficient sequence homogenization mechanisms, such as gene conversion or recurring unequal crossing-over, operate in T. vaginalis. The copy number of the Tv-E650 repeats was estimated to be about 10(2)-10(3) per genome. Based on the DNA hybridization results, the Tv-E650 repeat family is conserved in all T. vaginalis strains examined, regardless of their diverse geographical origin. No hybridization of the Tv-E650 probe was found with the DNA from Trichomonas tenax, Trichomonas gallinae and Pentatrichomonas hominis, indicating that the Tv-E650 repeated sequences are species-specific. A dot blot hybridization protocol was developed which does not require isolation of DNA. By using this protocol it was possible to detect the DNA released from approximately 10(3) T. vaginalis cells per dot. These observations suggest that the Tv-E650 probe is potentially applicable to the identification and detection of T. vaginalis.
从阴道毛滴虫基因组中克隆并测序了一个长度为650 bp的重复序列家族，命名为Tv-E650家族。核苷酸序列富含A+T（共有序列中A+T含量为73.3%），在所分析的8个分子克隆中高度保守。克隆之间的差异为单核苷酸和双核苷酸替换以及插入或缺失。克隆的序列一致性以及不同克隆中相同突变的存在表明，阴道毛滴虫中存在有效的序列均一化机制，如基因转换或反复出现的不等交换。估计每个基因组中Tv-E650重复序列的拷贝数约为10²-10³。基于DNA杂交结果，Tv-E650重复序列家族在所有检测的阴道毛滴虫菌株中均保守，无论其地理来源如何。未发现Tv-E650探针与口腔毛滴虫、鸡毛滴虫和人五毛滴虫的DNA杂交，表明Tv-E650重复序列具有种特异性。开发了一种无需分离DNA的斑点印迹杂交方法。使用该方法可以检测每个斑点中约10³个阴道毛滴虫细胞释放的DNA。这些观察结果表明，Tv-E650探针可能适用于阴道毛滴虫的鉴定和检测。
