【摘要】：目的構建穩(wěn)定過表達人表皮生長因子受體2(HER2/ErbB2)基因的黑素瘤細胞株,研究其對黑素瘤細胞的增殖、侵襲和遷移的影響。方法將pCMV3-ErbB2(人源)重組質粒通過脂質體法轉染至黑素瘤B16細胞株,用潮霉素B篩選出耐藥陽性克隆。實時定量PCR(qRT-PCR)檢測各單克隆ErbB2 mRNA相對表達量,免疫熒光技術檢測ErbB2蛋白的表達。MTT法檢測轉染后細胞增殖能力,TranswellTM實驗檢測細胞侵襲能力,劃痕實驗檢測細胞遷移能力。結果實時定量PCR檢測結果表明,通過脂質體法成功構建過表達HER2的B16細胞株,其HER2的表達顯著高于空載體對照細胞。TranswellTM等實驗顯示HER2過表達增強黑素瘤細胞增殖、侵襲和遷移的能力。結論過表達HER2促進黑素瘤細胞的增殖、侵襲和遷移。
[Abstract]:Objective to construct a melanoma cell line stably expressing human epidermal growth factor receptor 2 (HER2/ErbB2) gene and study its effect on the proliferation, invasion and migration of melanoma cells. Methods pCMV3-ErbB2 recombinant plasmid was transfected into melanoma B16 cell line by liposome method, and hygromycin B was used to screen the drug resistant clones. The relative expression of each monoclonal ErbB2 mRNA was detected by real-time quantitative PCR (qRT-PCR), the expression of ErbB2 protein was detected by immunofluorescence, the proliferative ability of transfected cells was detected by MTT assay, the ability of cell invasion was detected by TranswellTM assay, and the ability of cell migration was detected by scratch assay. Results the results of real-time quantitative PCR analysis showed that the expression of HER2 in B16 cell line which overexpressed HER2 was significantly higher than that in blank vector control cells by liposome method. TranswellTM and other experiments showed that HER2 overexpression enhanced the proliferation of melanoma cells. The ability to invade and migrate. Conclusion overexpression of HER2 promotes the proliferation, invasion and migration of melanoma cells.
【作者單位】： 新鄉(xiāng)醫(yī)學院生命科學技術學院河南省高校合成生物改造工程與應用重點實驗室;
【基金】：河南省重大科技專項研究課題(161100310900) 新鄉(xiāng)醫(yī)學院研究生科研創(chuàng)新支持計劃(YJSCX201634Y)
【分類號】：R739.5

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