摘要
为了早期快速诊断黄病毒感染，在其ＮＳ１基因序列设计了一对通用引物，扩增序列长度为４１３ｂｐ；在登革病毒（ＤＥＮ）１，２，３，４型和乙型脑炎病毒（ＪＥＶ）设计了各型的内引物，扩增序列长度分别是ＤＥＮ１型为２６２ｂｐ，ＤＥＮ２型为１８９ｂｐ，ＤＥＮ３型为３９２ｂｐ，ＤＥＮ４型为９７ｂｐ，ＪＥＶ为３２３ｂｐ。应用逆转录－聚合酶链反应（ＲＴ－ＰＣＲ）成功地扩增了ＤＥＮ１－４型和ＪＥＶ基因部分片段，其扩增片段的大小与设计相符合。应用套式ＰＣＲ检测临床诊断为登革热的患者血清标本７８份，证实ＤＥＮ１型阳性１８份，ＤＥＮ２型阳性４８份，其中８份同时合并感染ＤＥＮ４型。采用套式ＰＣＲ检测临床诊断为乙型脑炎患者血标本４２份，证实乙型脑炎病毒感染者３５份。结果表明，该法能直接检测发病患者早期血标本中的病毒基因，并在２天内完成对黄病毒的鉴别诊断。
For the purpose of early and rapid diagnosis of flavivirus infection, the universal primer set was selected on the NS1 gene. Another five different internal primers were selected on the NS1 genes of DEN 1,2,3,4 and JEV. With the universal primer set designed, the NS1 fragment about the size of 413bp and with the five internal primers, the NS1 fragments about the sizes of 262bp(DEN1), 189bp(DEN2), 392bp(DEN3), 97bp(DEN4) and 323bp(JEV) were amplified respectively. The reverse transcription polymerase chain reaction (RT-PCR) was used to amplify gene fragments of flaviviruses. cDNA of DEN1,2,3,4 and JEV were successfully amplified with universal primer set and internal primers. Sera from 78 patients with dengue fever were assayed by nested PCR, DEN1 was detected from 18 of the 40 patients' sera and DEN2 was detected from 48 patients' sera. Sera from 42 patients with Japanese encephalitis were also assayed by nested PCR, JEV was detected from 35 of the 42 patients' sera. By nested PCR, we completed identification of flaviviruses within 2 days. The results showed that this method has the potential value in rapid clinical diagnosis of flavivirus infection.
出处
《中华实验和临床病毒学杂志》
CAS
CSCD
1997年第3期267-270,共4页
Chinese Journal of Experimental and Clinical Virology
关键词
聚合酶链反应
黄病毒
登革热
病毒学
Polymerase chain reaction Flavivirus Dengue fever/Virology
