SIMULTANEOUS DETECTION OF HIV-1 AND HCV GENOMIC SEQUENCES BY PCR

建立了一种快速、简便的聚合酶链式反应(PCR)方法 为协同扩增和同时检测丙型肝炎而设计 病毒(丙型肝炎病毒)和人类免疫缺陷病毒1型(HIV-1)，特异性 同一血清样本中的序列。从13个样品中提取基因组RNA 同时检测抗-丙型肝炎病毒和抗-丙型肝炎病毒抗体阳性的献血员血清 HIV-1。将提取的rna反转录为cdna并进行扩增。 使用基于丙型肝炎病毒的嵌套引物对(SN01、SN02、SN03和SN04) 丙型肝炎病毒克隆37B和81，HIV-1克隆SK 38/39的原型序列 同时。聚合酶链式反应产物通过液体杂交和 32P末端标记寡核苷酸探针的Southern杂交 从两个引物对之间的区域，不包括引物区 序列。这些引物对能够检测到1-3个拷贝和 比较时，丙型肝炎病毒和艾滋病毒-1的基因组RNA分别为10-20个拷贝 以8E5细胞为单拷贝标准。13例患者均检出丙型肝炎病毒RNA (100%)检测样本；11(85%)样本检测到HIV-RNA。这个 从两个不同的病毒基因组中共扩增特定序列的能力 同样的反应混合物可以促进同时检测 多个病毒序列，从而提供了同时 在有症状的和非典型肺炎的血清标本中检测一种以上的病毒病原体 没有症状的个体。

A rapid and simple method using the polymerase chain reaction (PCR) was devised for the co-amplification and simultaneous detection of hepatitis C virus (HCV) and human immunodeficiency virus type 1 (HIV-1), specific sequences in the same serum sample. Genomic RNA was extracted from 13 blood donor sera that were reactive in ELISA for both anti-HCV and anti- HIV-1. The extracted RNA was reverse transcribed into cDNA and amplified using nested primer pairs (SN01, SN02, SN03 and SN04) based on the HCV prototype sequence of clones 37b and 81 for HCV, and SK 38/39 for HIV-1 simultaneously. PCR products were analyzed by liquid hybridization and Southern blot hybridization with 32P end-labeled oligonucleotide probes from the regions between the two primer pairs, excluding the primer sequences. The primer pairs were able to detect as few as 1-3 copies and 10-20 copies of genomic RNA for HCV and HIV-1, respectively, when compared with 8E5 cells as the single copy standard. HCV-RNA was detected in all 13 (100%) samples tested; HIV-RNA was detected in 11 (85%) samples. The ability to coamplify specific sequences from two different viral genomes in the same reaction mixture could facilitate the simultaneous detection of multiple viral sequences, thus offering the possibility of simultaneous diagnosis of more than one viral agent in serum samples of symptomatic and asymptomatic individuals.