革螨、恙螨体内HFRSV-RNA定位检测方法研究

摘要

The probers of hemorrhagic fever with renal syndrome virus(HFRSV)cDNA were labeled by Dig to detect HFRSV location in Gasmid mite and Chigger mite and the procedure of in situ hybridization were studied. The results showed that positive signals were detected in ovary cells and midgut cells. The effect were similar between gelatinchromium potassium sulfate and polylysine in pretreatment; sections were prevented from dropping efficiently room temperature 2h ,42℃1h and fixation in 4% paraformaldehyde and kept well structure. Background would be light while sperm DNA and yeast tRNA were added in prehybridization. Optional results would come up after hybridized 16h in 42℃ and color developed 7h in color solution.%用地高辛素标记肾综合征出血热病毒cDNA分子探针，检测肾综合征出血热病毒在革螨、恙螨体内的分布定位，并对原位分子杂交的步骤进行研究。结果表明：病毒核酸信号呈弥散分布，主要位于卵巢细胞，中肠及支囊细胞。在铺片时，明胶-硫酸铬钾与多聚赖氨酸的效果相似，4%多聚甲醛的固定效果较好，可有效防止脱片。预杂交中加入SpermDNA和yeasttRNA，可有效地减少背景染色；42℃杂交16h，显色7h，可得到理想结果。