【摘要】 蓖麻蚕核糖体RNA基因非转录间隔区 (NTS)有一长约 1kb的核骨架结合区 (scaffold associatedregion ,SAR)。将这一序列克隆到含荧光素酶 (luciferase)报告基因的真核表达载体 pLu中 ,在阳离子脂质体的介导下对NIH3T3细胞进行瞬时转染 (transienttransfection)及稳定转化 (stabletransformed)株的筛选 ,然后检测SAR在瞬时转染及稳定整合条件下对荧光素酶报告基因表达活性的影响。实验结果表明 ,在稳定转化的细胞中 ,该SAR对luciferase基因的表达有明显的激活作用 ,增强基因表达活性约 15倍 ;而在瞬时表达的条件下 ,SAR激活基因表达的作用不明显。说明SAR增强基因的表达可能需要整合在染色体上才能完成。我们用Southwestern印迹法实验进一步证实了蓖麻蚕rRNA基因的SAR能特异性地与NIH3T3细胞核骨架蛋白结合 ,因此该SAR的作用具有非组织特异性更多还原

【Abstract】 The 1 kb scaffold attachment region (SAR) at 5′ non transcription region of rRNA gene of silkworm Attacus ricini was cloned into eukaryotic expression vector pLu, which contained luciferase report gene and neo R selecting marker. After transfection of constructs into NIH3T3 cell line by using cation liposome, the luciferase activity was monitored to check the SAR’s function. The results demonstrated that the SAR could enhance gene expression up to 15 fold in stable transformed cells, but no obvious gene expression was observed in transient transfection. Its effect on gene expression appeared to require chromosomal integration. Southwestern blotting experiments showed that SAR specifically bound to nuclear matrix proteins of NIH3T3 cells. The binding with nuclear matrix may be necessary for SAR function of transcriptional enhancement.更多还原