Direct detection of HIV-1 RNA from AIDS and ARC patient samples.
Direct detection of HIV-1 RNA from AIDS and ARC patient samples.
复制标题
直接检测 AIDS 和 ARC 患者样本中的 HIV-1 RNA。
DOI：
10.1089/dna.1988.7.287
复制
发表时间：
1988
期刊：
影响因子：
--
通讯作者：
J. Rossi
中科院分区：
文献类型：
--
作者：
G. Murakawa;J. Zaia;P. Spallone;DELILAH A. Stephens;B. Kaplan;R. Wallace;J. Rossi
人类免疫缺陷病毒（Human immunodeficiency virus，HIV），又称人类嗜T淋巴细胞病毒（human T-lymphotropic virus，HTLVIII/LAV），是获得性免疫缺陷综合征（acquired immune deficiency syndrome，AIDS）的病原体。由于感染细胞的百分比低，仅在一小部分病例中可以直接检测患者组织或血液样品中的HIV-1核酸序列（Shaw等人，1984年）。我们报道了聚合酶链反应方法（PCR）的改进（Saiki等人，1985），其中我们从HIV-1 RNA模板扩增序列，用于鉴定从AIDS和AIDS相关复合体（ARC）患者获得的外周血和组织样品中的HIV-1。这种HIV-1检测方法比标准核酸检测方法灵敏至少六个数量级，具有直接的临床应用价值。HIV-1检测不需要进行病毒的体外组织培养。使用这种技术，在HIV-1的orfB区域的序列已被扩增，并从少于1微克的总RNA从几毫升的外周血样品制备检测。该技术能够快速和明确的临床检测潜在的HIV感染者，并可用于测定抗HIV-1药物的疗效。为了提高该技术的效率，我们将原核T7 RNA聚合酶启动子序列附加到其中一个引发寡核苷酸上。在用含启动子的寡核苷酸进行几个PCR循环后，可以利用反应的一小部分来指导特异性和有效的T7 RNA聚合酶介导的扩增序列的转录，从而提高灵敏度并简化实验的劳动。
Human immunodeficiency virus (HIV), formerly termed human T-lymphotropic virus (HTLVIII/LAV), is the etiological agent of acquired immune deficiency syndrome (AIDS). Direct detection of HIV-1 nucleic acid sequences in patient tissue or blood samples is possible in only a minor fraction of cases due to the low percentage of infected cells (Shaw et al., 1984). We report a modification of the polymerase chain reaction method (PCR) (Saiki et al., 1985), in which we amplify sequences from HIV-1 RNA templates, for the identification of HIV-1 in peripheral blood and tissue samples obtained from AIDS and AIDS-related complex (ARC) patients. This method of HIV-1 detection is at least six orders of magnitude more sensitive than standard nucleic acid detection methods and has direct clinical applications. In vitro tissue culturing of the virus is not required for HIV-1 detection. Using this technique, the sequence in the orfB region of HIV-1 has been amplified and detected from less than 1 microgram of total RNA prepared from a few milliliters of peripheral blood samples. This technique enables the rapid and unambiguous clinical detection of potential HIV-infected individuals and can be used to assay the efficacy of anti-HIV-1 drugs. To enhance the efficiency of this technique, we have appended the prokaryotic T7 RNA polymerase promoter sequence to one of the priming oligonucleotides. After several cycles of PCR with the promoter-containing oligo, a small aliquot of the reaction can be utilized to direct specific and efficient T7 RNA polymerase-mediated transcription of the amplified sequences, thus enhancing the sensitivity and simplifying the labor of the experiment.
人类免疫缺陷病毒突变分析：orf-B区下调病毒复制。
DOI：
10.1073/pnas.84.5.1434
发表时间：
1987
影响因子：
11.1
作者：
Luciw,PA;Cheng-Mayer,C;Levy,JA
通讯作者：
Levy,JA