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目的:设计并构建ErbB2的小干扰RNA,检测其对ErbB2蛋白表达的干扰效果,并检测其对乳腺癌细胞ZR75-1生长的影响。方法:设计2条针对ErbB2的siRNA,并克隆到siRNA表达载体pSliencer 2.1-U6 neo上。经酶切和测序证明构建成功后,将其与pcDNA3-FLAG-ErbB2共转染于293T细胞,以及单转siRNA于乳腺癌SKBR3和ZR75-1细胞,通过Western blot分别检测siRNA对外源和内源ErbB2的干扰效果。通过结晶紫实验研究siRNA对ZR75-1生长的影响。结果:Western blot证明构建的两条ErbB2 siRNA均能有效抑制外源和内源ErbB2蛋白的表达,并抑制乳腺癌ZR75-1细胞的生长。结论:构建的siRNA能有效地抑制ErbB2蛋白的表达并抑制ZR75-1细胞的生长。
OBJECTIVE: To design and construct ErbB2 small interfering RNA (RNAi) to detect its interference effect on ErbB2 protein expression and to detect its effect on the growth of breast cancer cell line ZR75-1. Methods: Two siRNAs targeting ErbB2 were designed and cloned into siRNA expression vector pSliencer 2.1-U6 neo. The recombinant plasmid pcDNA3-FLAG-ErbB2 was co-transfected into 293T cells and single-transfection siRNA into SKBR3 and ZR75-1 cells. Western blot was used to detect the effect of siRNA on exogenous and endogenous ErbB2 interference effect. The effect of siRNA on the growth of ZR75-1 was studied by crystal violet experiment. Results: Western blot showed that both ErbB2 siRNAs could effectively inhibit the expression of ErbB2 protein and inhibit the growth of breast cancer cell line ZR75-1. Conclusion: The constructed siRNA can effectively inhibit the expression of ErbB2 protein and inhibit the growth of ZR75-1 cells.