萤火虫荧光素酶基困的反义RNA及其缺失片段阻抑基因表达的研究

Studies on the Antisense RNA of Firefly Luciferase Gene and the Blocknig of Gene Expression by the RNA Deletion Segments of the Luciferase Gene

利用离体转录载体ｐＳＰ１８、ｐＧＥＭ－３Ｚ和萤火虫荧光素酶基因（Ｌｕｃ）及其缺失片段构建了一组融合质粒。以这些质粒为模板，通过离体转录产生荧光素酶基因的ｍＲＮＡ，反义ＲＮＡ及大小和结构不同的反义ＲＮＡ缺失片段，再通过显微注射分别将其引入爪赡卵母细胞的细胞质。注射后的卵母细胞经一定时间培育，然后检测其翻译产物一荧光素酶的相对活性。结果表明，荧光素酶基因的表达受反义ＲＮＡ及其缺失片段的严重阻抑，表达抑制和反义ＲＮＡ之间存在明显的剂量效应。而且，基因表达的抑制程度主要的和反义ＲＮＡ片段的结构有关。

In vitro transcriptional vectors' pSP18, PGEM-3Z and the cDNA gene of the firefly luciferase were used to construct a set of fusion plasmids.These plasmids were used as templates and yielded a mRNA of the firefly luciferase gene, its antisense RNA and a series of deletion fragments of the antisense RNA with different sizes and structures by transcription in vitro.Then, they were introduced into the cytoplasm of Xenopus oocytes by microinjection , respectively. The injected oocytes were incubated for definite periods and assayed their relative activities of the luciferase. Results suggested that the gene expression of the firefly luciferase are inhibited seriously by the antisense RNA and its deletion fragments. And there is a marked dosage effect between the inhibition of gene expression and amounts of the antisense RNA. In addition,the inhibition levels of the gene expression are majorly dependent on the structures of the antisense RNA.