靶向Rab9 GTPase siRNA表达载体的构建和鉴定

目的构建靶向Rab 9 GTPase siRNA表达载体。方法设计有小发夹RNA结构靶向Rab9GT-Pase的68个碱基的寡核苷酸,克隆入表达载体pSUPER.neo EGFP,得到重组质粒pSUPER.neo EGFP-R1和pSUPER.neo EGFP-R2,通过限制性核酸内切酶酶切和DNA序列测定对重组质粒进行鉴定。结果经酶切鉴定寡核苷酸已成功插入表达载体pSUPER.neo EGFP,DNA序列分析表明插入片段序列与合成的siR-NA序列相同。结论成功构建了靶向Rab9 GTPase siRNA表达载体,为进一步研究Rab9 GTPase与麻疹病毒复制之间的关系奠定基础。

Construction and identification of siRNA expression vector targeting for Rab9 GTPase

Objective] To construct small interference RNA (siRNA) expression vector targeting for Rab9 GTPase. Methods] Oligos of 68 base pairs for hairpin RNA targeting for Rab9 GTPase were designed and cloned into the expression vector pSUPER. neo EGFP to generate the recombinant plasmid pSUPER. neo EGFP-R1 and pSUPER. neo EGFP-R2. The recombinant plasmid was confirmed with restriction endonuclease digestion and DNA sequencing. Results] Expression vector was constructed successfully and identified by enzyme digestion analysis. DNA sequence analysis of inserted fragment revealed the same sequences as synthesized siRNA oligonucleotides. Conclusion] The Rab9 GTPase specific small interfering RNA expression vector has been constructed successfully which facilitates the study on relationship between Rab9 GTPase and replication of measles virus.