抑制绿色荧光蛋白的慢病毒表达载体的构建

Construction of a lentiviral expression vector that inhibits enhanced green fluorescent proteins

目的:构建抑制增强型绿色荧光蛋白(EGFP)表达的慢病毒表达载体。方法:选取EGFP的19nt特异性序列,构建针对EGFP的shRNA,用XbaⅠ、XhoⅠ双酶切后定向克隆到慢病毒表达载体pLL3.7中,用包装的慢病毒液感染细胞,用荧光显微镜观察EGFP基因在不同细胞的表达情况。结果:干扰序列正确克隆至含U6启动子的慢病毒载体中,慢病毒感染Hela及J558L细胞后,可有效抑制细胞中EGFP的表达。结论:构建的慢病毒系统有效抑制了EGFP的表达,为后续的基因功能研究提供了有利工具。

Objective ： To construct a lentiviral expression vector that inhibits enhanced green fluorescent proteins （EGFP）. Methods： The EGFP shRNA was cloned into the pLL3.7 lentivirus vector digested with Xba I/Xho I . J558L and Hela cells were infected by lentiviruses, and the EGFP expression was observed with the fluorescence microscope. Results： The EGFP RNAi sequence was correctly cloned into the lentivirus vector with the U6 promoter, and the EGFP expressions of Hela and J558L cells were suppressed after infected by lentivirus containing EGFP shRNA. Conclusion ： The lentivirus vector effectively inhibited EGFP expression, which could be used to mediate further researches on gene function.