Minimum protein requirements for transcription and RNA replication of a minigenome of human parainfluenza virus type 3 and evaluation of the rule of six
Minimum protein requirements for transcription and RNA replication of a minigenome of human parainfluenza virus type 3 and evaluation of the rule of six
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DOI：
10.1006/viro.1997.8633
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发表时间：
1997-07-21
期刊：
影响因子：
3.7
通讯作者：
Collins, PL
中科院分区：
文献类型：
--
作者：
Durbin, AP;Siew, JW;Collins, PL
建立了人副流感病毒3型（HPIV 3）的重组转录和RNA复制系统。该系统基于HPIV 3基因组RNA的反义类似物，其中病毒基因被删除并被编码细菌氯霉素乙酰转移酶（CAT）的基因所取代。从共转染质粒表达的N、P和L蛋白对于指导有效的转录和RNA复制是必要的和足够的。转录产生亚基因组聚腺苷酸化的mRNA，其通过寡聚（dT）色谱分离。RNA复制产生了一个小的反基因组和后代的小基因组，这被证明是基于微球菌核酸酶消化的抗性。构建一组cDNA以编码长度相差单核苷酸增量的微型基因组。重组系统中的转录和RNA复制对于长度为6的偶数倍的微型基因组是最有效的。RNA的复制和转录似乎都受这一规律的支配。然而，长度大于或小于6的偶数倍的一个核苷酸的微型基因组也非常活跃，特别是在RNA复制中，这表明规则不是绝对的。(C)北京：科学出版社.
A reconstituted transcription and RNA replication system for human parainfluenza virus type 3 (HPIV3) was developed using components expressed intracellularly from transfected plasmids driven by T7 RNA polymerase supplied by a vaccinia virus recombinant The system is based on a negative-sense analog of HPIV3 genomic RNA in which the viral genes were deleted and replaced with that encoding bacterial chloramphenicol acetyl transferase (CAT). The N, P, and L proteins expressed from cotransfected plasmids were necessary and sufficient to direct efficient transcription and RNA replication. Transcription yielded subgenomic polyadenylated mRNA, which was isolated by oligo(dT) chromatography. RNA replication yielded a mini-antigenome and progeny minigenome, which were shown to be encapsidated based on resistance to digestion with micrococcal nuclease. A panel of cDNAs was constructed to encode minigenomes which differed in length by single-nucleotide increments. Transcription and RNA replication in the reconstituted system were most efficient for the minigenome whose length was an even multiple of six. Both RNA replication and transcription appeared to be governed by the rule. However, minigenomes whose lengths were one nucleotide greater than or less than an even multiple of six also were very active, especially in RNA replication, indicating that the rule was not absolute. (C) 1997 Academic Press.