Liu D X, Inglis S C
Department of Pathology, University of Cambridge, United Kingdom.
Virology. 1992 Jan;186(1):342-7. doi: 10.1016/0042-6822(92)90094-6.
The second smallest subgenomic messenger RNA, mRNA5, of the coronavirus infectious bronchitis virus includes in its "5' unique region" two separate open reading frames (5a and 5b), whose coding function has not so far been established, and thus it may represent a dicistronic messenger RNA. We report here that two polypeptides with the sizes expected for the 5a and 5b products can be synthesised by in vitro translation of a single artificial mRNA containing both the 5a and 5b ORFs. To establish whether these polypeptides represent genuine virus gene products, both the 5a and 5b coding sequences were expressed as bacterial fusion proteins, and these were used to raise monospecific antisera. Antisera raised against both the 5a and 5b-specific sequences recognized specifically proteins of the expected size in infectious bronchitis virus-infected chicken kidney and Vero cells, indicating that 5a and 5b do represent genuine virus genes, and suggesting that mRNA5 is indeed functionally dicistronic.
传染性支气管炎病毒是一种冠状病毒，其第二小的亚基因组信使核糖核酸（mRNA5）在其“5' 非编码区”包含两个独立的开放阅读框（5a和5b），其编码功能迄今尚未明确，因此它可能代表一种双顺反子信使核糖核酸。我们在此报告，通过对包含5a和5b开放阅读框的单一人工信使核糖核酸进行体外翻译，可合成两种大小符合5a和5b产物预期的多肽。为确定这些多肽是否代表真正的病毒基因产物，5a和5b编码序列均被表达为细菌融合蛋白，并用于制备单特异性抗血清。针对5a和5b特异性序列产生的抗血清在传染性支气管炎病毒感染的鸡肾细胞和非洲绿猴肾细胞（Vero细胞）中特异性识别了预期大小的蛋白质，这表明5a和5b确实代表真正的病毒基因，并提示mRNA5在功能上确实是双顺反子。
