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目的:针对DREAM基因中外显子序列,设计并筛选出起作用的siRNA,为进一步研究以DREAM为靶标的基因治疗提供依据。方法:通过利用生物信息学的方法设计出66条潜在的针对DREAM基因中外显子序列的siRNA序列。潜在的序列根据G+C含量分析及Genbank BLAST检测,我们筛选了三条较理想的DREAM基因siRNA靶标,并将其构建到pENTR/H1/TO载体中,转染细胞后提取总蛋白并用Western blot方法检测DREAM蛋白的表达水平。结果:示外显子9中5’末端位置为1253的序列能够抑制80%的DREAM的蛋白表达。结论:这一研究结果为进一步siRNA类药物的实验研究提供了理论基础。
OBJECTIVE: To design and screen siRNAs that target exon sequences in DREAM gene and provide basis for further study of gene therapy targeting DREAM. METHODS: Sixty-six siRNA sequences targeting exon sequences in DREAM gene were designed by bioinformatics methods. According to the G + C content analysis and the GenBank BLAST assay, we screened three more ideal siRNA targets of DREAM gene and constructed it into pENTR / H1 / TO vector. After transfection, the total protein was extracted and analyzed by Western blot The expression of DREAM protein was detected. Results: The sequence showing 1253 at the 5 ’end of exon 9 was able to inhibit the protein expression of 80% of DREAM. Conclusion: This study provides the theoretical basis for further experimental study of siRNAs.