SENSITIVE DNA HYBRIDIZATION TEST FOR ROTAVIRUS RNA

轮状病毒是婴儿严重腹泻的主要原因。这 年确诊的胃肠炎病例中病原体占40%-60% 两岁以下的儿童。目前诊断是通过酶联免疫吸附试验进行的 或者通过电子显微镜观察。后者对于小企业来说是不切实际的。 实验室，而ELISA产生了令人无法接受的假数量 积极的一面。该病毒由11个双链RNA片段组成。这个 主要衣壳糖蛋白基因的核苷酸序列已知 三种人类血清型。我们建议合成DNA探针，这将 与人类共有的基因组区域特异杂交 血清型。 第一阶段，根据已公布的序列信息，我们将合成一个 与病毒正链RNA互补的寡核苷酸探针 基因组和病毒的信使核糖核酸。探测器将接受能力测试 检测典型粪便样本中的轮状病毒。主要技术 这项工作的创新将是开发能够 用于检测双链RNA病毒的寡核苷酸探针 临床样本。 第二阶段。我们将对合成的最好的探针进行临床试验， 主要是非同位素标记的，以产生临床 快速检测粪便样本中轮状病毒的诊断试剂盒。

Rotaviruses are the major cause of serious infantile diarrhea. This pathogen accounts for 40-60% of the diagnosed cases of gastroenteritis in children under the age of two years. Diagnosis is currently made by ELISA or by electron microscopy. The latter is impractical for the small laboratory, and the ELISA produces an unacceptable number of false positives. The virus consists of 11 segments of double-stranded RNA. The nucleotide sequence of the major outer capsid glycoprotein gene is known for three human serotypes. We propose to synthesize DNA probes which will hybridize specifically with a region of the genome common to the human serotypes. Phase I. Based on published sequence information, we will synthesize an oligonucleotide probe complementary both to the plus strand RNA of the genome and the mRNA of the virus. The probe will be tested for its ability to detect rotavirus in characterized stool samples. The major technical innovation in this work will be the development of methods which allow oligonucleotide probes to be used to detect double-stranded RNA viruses in clinical samples. Phase II. We will conduct clinical trials of the best probes synthesized, primarily non-isotopically labeled in order to produce a clinical diagnostic kit to rapidly detect rotavirus in stool samples.