Deletion of an immunodominant Trypanosoma cruzi surface glycoprotein disrupts flagellum-cell adhesion.
Deletion of an immunodominant Trypanosoma cruzi surface glycoprotein disrupts flagellum-cell adhesion.
复制标题
免疫主导锥虫表面糖蛋白的删除破坏鞭毛 - 细胞粘附。
DOI：
10.1083/jcb.122.1.149
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发表时间：
1993-07
期刊：
影响因子：
--
通讯作者：
Cross GA
中科院分区：
文献类型：
--
作者：
Cooper R;de Jesus AR;Cross GA
克氏锥虫昆虫阶段特异性糖蛋白 GP72 的无效突变体是通过靶向基因替换产生的。构建了靶向质粒，其中新霉素磷酸转移酶和潮霉素磷酸转移酶基因的侧翼为GP72序列。通过电穿孔将这些质粒依次转染到克氏锥虫上鞭毛体中。 Southern印迹分析表明这两个基因发生了精确替换。 GP72 基因座没有发生异常重排，并且没有 GP72 基因序列易位到基因组的其他位置。蛋白质印迹证实 GP72 在这些无效突变体中不表达。突变体的形态与野生型显着不同。在突变型和野生型寄生虫中，鞭毛从鞭毛袋中出现。在无效突变体中，鞭毛与寄生虫细胞膜的正常附着丧失。
Null mutants of the Trypanosoma cruzi insect stage-specific glycoprotein GP72 were created by targeted gene replacement. Targeting plasmids were constructed in which the neomycin phosphotransferase and hygromycin phosphotransferase genes were flanked by GP72 sequences. These plasmids were sequentially transfected into T. cruzi epimastigotes by electroporation. Southern blot analyzes indicated that precise replacement of the two genes had occurred. No aberrant rearrangements occurred at the GP72 locus and no GP72 gene sequences had been translocated elsewhere in the genome. Western blots confirmed that GP72 is not expressed in these null mutants. The morphology of the mutants is dramatically different from wild-type. In both mutant and wild-type parasites, the flagellum emerges from the flagellar pocket. In the null mutant the normal attachment of the flagellum to the cell membrane of the parasite is lost.