Fordham-Skelton A P, Taylor P N, Hartley M R, Croy R R
Department of Biological Sciences, University of Durham, UK.
Mol Gen Genet. 1991 Oct;229(3):460-6. doi: 10.1007/BF00267470.
A. Saponaria (soapwort) genomic library was screened with a PCR-derived saporin-specific gene probe. The nucleotide sequences of three saporin genomic clones were determined. One of the clones contained a full-length saporin coding sequence whilst the other two were truncated. A hybrid full-length saporin coding sequence was constructed using the two truncated clones. An SP6 promoter sequence and in-frame initiation codon was added to each of the coding sequences using PCR. In vitro translation of saporin coding sequence transcripts in rabbit reticulocyte lysates resulted in the specific depurination of 28S RNA. This indicated that the saporin sequences encoded functional polypeptides with RNA N-glycosidase activity.
A. 用PCR衍生的皂草素特异性基因探针筛选肥皂草基因组文库。测定了三个皂草素基因组克隆的核苷酸序列。其中一个克隆包含全长皂草素编码序列,而另外两个是截短的。使用这两个截短的克隆构建了一个杂交全长皂草素编码序列。通过PCR向每个编码序列中添加了一个SP6启动子序列和读框内起始密码子。在兔网织红细胞裂解物中对皂草素编码序列转录本进行体外翻译,导致28S RNA发生特异性脱嘌呤作用。这表明皂草素序列编码具有RNA N-糖苷酶活性的功能性多肽。