DEVELOPMENT OF UNIVERSAL PCR ASSAY FOR RETROVIRUSES

逆转录病毒通用 PCR 检测方法的开发

• 批准号: 3362264
• 负责人: EDOUARD M CANTIN
• 金额: $ 25.86万
• 依托单位: CITY OF HOPE NATIONAL MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-08-01 至 1992-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3362264
• 关键词: AIDS /HIV test; RNA directed DNA polymerase; Retroviridae; affinity labeling; diagnosis design /evaluation; fluorescent dye /probe; human T cell lymphotropic virus type 1; human T cell lymphotropic virus type 2; human immunodeficiency virus 1; human immunodeficiency virus 2; human tissue; molecular cloning; natural gene amplification; nucleic acid chemical synthesis; nucleic acid probes; nucleic acid sequence; virus DNA; virus classification; virus genetics

Retroviruses are ubiquitous in nature and have long been recognized as infectious cancer causing pathogens in animals. It was only in the last decade, however, that pathogenic human retroviruses were discovered. The human T lymphotropic viruses types I and II (HTLV 1 and HTLV 2) are causally associated with adult T-cell leukemia/lymphoma, while the human immunodeficiency viruses types I and II (HIV 1 and HIV 2) are the etiologic agents of AIDS - the modern plague. Clearly, these and other as yet uncharacterized retroviruses represent a considerable public health threat because their prevalence in the population appears to be increasing, as is their association with various diseases. Of particular concern is the fact that they are transmitted by contaminated blood and blood products, necessitating the implementation of screening procedures to ensure safety of the blood supply. Recent advances in molecular biology, such as the polymerase chain reaction (PCR), which facilitates in vitro amplification of DNA, create exciting new possibilities for DNA diagnostics. Our proposal has two major goals; the first is development of a PCR assay to amplify pol gene sequences in all known, as well as uncharacterized human retroviruses. This is feasible because the PCR primers are targeted to highly conserved regions of the pol gene; the second major goal is to automate the entire PCR assay to detect retroviruses, including the non-radioactive detection of the specifically amplified pol gene fragment. The long term goal is to perfect the assay to the point where it can be used in blood banks as a reliable, sensitive, simple, automated test for retrovirus contamination of blood and related products.

逆转录病毒在自然界中普遍存在，长期以来一直被认为是 动物体内引起传染性癌症的病原体。 这只是在最后 然而，十年后，致病性人类逆转录病毒被发现。 这 人类 T 淋巴细胞病毒 I 型和 II 型（HTLV 1 和 HTLV 2） 与成人 T 细胞白血病/淋巴瘤有因果关系，而人类 I 型和 II 型免疫缺陷病毒（HIV 1 和 HIV 2）是病因 艾滋病的病原体——现代瘟疫。显然，这些和其他的 未表征的逆转录病毒对公共卫生构成相当大的威胁 因为它们在人群中的患病率似乎在增加 它们与各种疾病的关联。特别值得关注的是一个事实 它们是通过受污染的血液和血液制品传播的， 需要实施筛查程序以确保安全 的血液供应。分子生物学的最新进展，例如 聚合酶链式反应 (PCR)，有利于体外扩增 DNA，为 DNA 诊断创造令人兴奋的新可能性。我们的建议 有两个主要目标；第一个是开发 PCR 检测来放大 pol 所有已知的以及未表征的人类逆转录病毒的基因序列。 这是可行的，因为 PCR 引物针对的是高度保守的 pol基因的区域；第二个主要目标是使整个过程自动化 PCR 检测逆转录病毒，包括非放射性检测 特异性扩增的 pol 基因片段。长期目标是 完善该检测方法，使其可以在血库中用作 可靠、灵敏、简单、自动化的逆转录病毒污染检测 血液及相关产品。