Smith D F, Searle S, Ready P D, Gramiccia M, Ben-Ismail R
Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, U.K.
Mol Biochem Parasitol. 1989 Dec;37(2):213-23. doi: 10.1016/0166-6851(89)90153-9.
A restriction fragment from a cloned kinetoplast minicircle DNA has been shown to be diagnostic for Leishmania major. This 402-bp TaqI fragment has been used routinely (as a radiolabelled probe) to detect 10(4) parasites in simple dot blots, both experimentally and in epidemiological surveys. It positively identified all stocks of L. major tested (including all six known zymodemes) and showed very low homology to kinetoplast DNA (kDNA) and chromosomal DNA of Leishmania infantum and Leishmania tropica, two species commonly isolated from patients and wild hosts within foci of L. major in the Old World. DNA sequence analysis of a minicircle of L. major is reported for the first time, and it is demonstrated that this species shares with Leishmania aethiopica, Sauroleishmania tarentolae and several species of Trypanosoma a region of conserved sequence that is involved in DNA replication, a process that could present targets for selective chemotherapeutic attack. Sequence and restriction fragment analyses have indicated the difficulties of selecting species-specific sequences from kDNA which, even in the same parasite clone, contains several predominant minicircle classes, not all of which contain diagnostic sequences.
已证明来自克隆的动基体小环DNA的一个限制性片段可用于诊断硕大利什曼原虫。这个402碱基对的TaqI片段已被常规用作（放射性标记探针），在实验和流行病学调查中，在简单的斑点印迹中检测10⁴个寄生虫。它能准确鉴定所有测试的硕大利什曼原虫毒株（包括所有六个已知酶解模式），并且与婴儿利什曼原虫和热带利什曼原虫的动基体DNA（kDNA）及染色体DNA的同源性非常低，这两种利什曼原虫是在旧大陆硕大利什曼原虫疫源地内常见于患者和野生宿主的物种。首次报道了硕大利什曼原虫一个小环的DNA序列分析，结果表明该物种与埃塞俄比亚利什曼原虫、蜥蜴利什曼原虫及几种锥虫属物种共享一段参与DNA复制的保守序列区域，而DNA复制过程可能成为选择性化疗攻击的靶点。序列和限制性片段分析表明，从动基体DNA中选择物种特异性序列存在困难，因为即使在同一个寄生虫克隆中，动基体DNA也包含几个主要的小环类别，并非所有类别都含有诊断序列。
