Solís F, Orozco E
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados I.P.N., Mexico, D.F., Mexico.
Exp Parasitol. 1991 Apr;72(3):236-42. doi: 10.1016/0014-4894(91)90142-j.
Transference of DNA to Entamoeba histolytica was carried out by polyethylene glycol fusion of two amebic clones with different phenotypes. Clone C9, strain HM1:IMSS, was the donor. It is emetine-resistant, highly phagocytic and virulent, and grows in soft agar. Clone L6, strain HM1:IMSS, the recipient, is emetine-sensitive, phagocytic, and virulence-deficient, and it does not grow in soft agar. Clones L6 and C9 have shown high stability in their virulence phenotypes since their isolation more than 5 years ago. Before fusion experiments, clone C9 was incubated in 20 micrograms/ml bromodeoxyuridine for 24 hr, and then, irradiated with 310 nm light to complete inactivation. Controls ensured that all irradiated trophozoites died after 24 hr of incubation at 37 degrees C. Irradiated C9 trophozoites were fused with L6 trophozoites, and hybrids were selected by their ability to grow in the presence of emetine. All hybrids, independently generated, grew poorly in soft agar and showed both an intermediate emetine-resistance and rate of phagocytosis. Some of them destroyed efficiently cell monolayers, but interestingly, they showed differences in their ability to produce hepatic abscesses in hamsters.
通过聚乙二醇融合两个具有不同表型的阿米巴克隆，将DNA转移至溶组织内阿米巴。供体是克隆C9，菌株HM1:IMSS。它对依米丁耐药，具有高度吞噬性和毒性，且能在软琼脂中生长。受体是克隆L6，菌株HM1:IMSS，对依米丁敏感，具有吞噬性，但缺乏毒性，且不能在软琼脂中生长。自5年多前分离以来，克隆L6和C9的毒力表型显示出高度稳定性。在融合实验前，将克隆C9在20微克/毫升溴脱氧尿苷中孵育24小时，然后用310纳米的光照射以完成灭活。对照确保所有经照射的滋养体在37摄氏度孵育24小时后死亡。将经照射的C9滋养体与L6滋养体融合，并通过它们在依米丁存在下生长的能力来选择杂种。所有独立产生的杂种在软琼脂中生长不良，显示出中等程度的依米丁耐药性和吞噬率。其中一些能有效地破坏细胞单层，但有趣的是，它们在仓鼠体内产生肝脓肿的能力存在差异。
