RIBOZYMES THAT CLEAVE POTATO LEAFROLL VIRUS-RNA WITHIN THE COAT PROTEIN AND POLYMERASE GENES

被引:18
作者
LAMB, JW [1 ]
HAY, RT [1 ]
机构
[1] UNIV ST ANDREWS,DEPT BIOCHEM & MICROBIOL,IRVINE BLDG,N STREET,ST ANDREWS KY16 9AL,FIFE,SCOTLAND
关键词
D O I
10.1099/0022-1317-71-10-2257
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
Two ribozymes were synthesized which were designed to cleave potato leafroll virus (PLRV) positive strand RNA within the regions known to encode the viral coat protein and the predicted RNA polymerase gene. DNA sequences encoding the ribozymes were inserted into the Escherichia coli plasmids pTz18R and pTz19R under the control of the bacteriophage T7 promoter and enzymically active RNA molecules generated by transcription by T7 RNA polymerase in vitro. Each ribozyme cleaved its cognate site in RNA derived from either cDNA or PLRV particles. Ribozyme cleavage sites within the polymerase gene and coat protein gene were determined and shown to be at the predicted sequence immediately downstream from a GUC motif. An altered version of the ribozyme which recognized the sequence in the coat protein gene was isolated in which a single adenosine residue in the enzymic loop of the ribozyme was deleted. This mutated ribozyme was unable to cleave RNA molecules containing the coat protein ribozyme target site.
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页码:2257 / 2264
页数:8
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