BIOLOGY OF NON-A, NON-B HEPATITIS AGENTS

迈克尔·霍顿和他在CHIRON公司的同事克隆了， NANBH至少部分基因组的表达和测序 探员。他们认为这是一个10kb的正义多聚腺苷 可能与阿尔法病毒有亲缘关系的RNA病毒。数列 在NANBH部分序列和NANBH部分序列之间没有发现同源性 任何其他已知的病毒序列。我计划用多种不同的cDNA 克隆程序，以确认CHIRON的发现。 然而，由于他们一开始是用公升的血浆 含有106个传染性病毒/毫升，我有大约100个 一毫升相同效价的血浆，任务很可能非常 很难。希望聚合酶链式反应 技术将有助于这一努力。 非甲非乙型肝炎的分子鉴定方法 代理现在涉及到技术的开发，可以用于 克隆极少量的核酸。我们知道， NANBH试剂在血清甚至肝脏中的含量都很低， 即每毫升血清或每克血清少于106个感染单位 肝脏。此外，似乎如果产生Poly(A)+NANBH mRNA，它 是一个非常低丰度的信息。因此，要么直接克隆 从技术上讲，克隆DNA或cDNA是非常困难的。为了 协助克隆少量DNA或cDNAs，一种“不能克隆” 已经产生了一段RNA或DNA。这个不可克隆的片段 将添加到NANBH核酸制剂中作为 所有克隆步骤的载体。这个不可克隆的片段将是 适用于多种不同的克隆策略，其中 感兴趣的核酸的含量非常低。

Michael Houghton and colleagues at Chiron Corporation have cloned, expressed and sequenced at least part of the genome of the NANBH agent. They believe it is a 10KB, positive sense polyadenylated RNA virus that may be related to the alpha togaviruses. Sequence homology has not been found between the NANBH partial sequence and any other known viral sequence. I plan to use a variety of cDNA cloning procedures in order to confirm the Chiron findings. However, since they started with liter quantities of plasma that contained 106 infectious viruses/ml and I have approximately 100 ml of plasma of the same titer, the task is likely to be very difficult. It is hoped that the polymerase chain reaction technique will aid this effort. Molecular approaches to identifying the non-A, non-B hepatitis agent now involve development of technology that can be used to clone extremely small quantities of nucleic acid. We know that the NANBH agent is present in serum and even liver at very low levels, i.e., less than 106 infectious units per ml of serum or gram of liver. It also seems that if poly(A)+NANBH mRNA is produced, it is a very low abundance message. Therefore, either direct cloning of DNA or cDNA cloning is technically very difficult. In order to assist cloning small quantities of DNA or cDNA, a "nonclonable" piece of RNA or DNA has been produced. This non-clonable fragment will be added to NANBH nucleic acid preparations to act as a carrier for all cloning steps. This non-clonable fragment will be useful for a number of different cloning strategies where the level of the nucleic acid of interest is very low.