GATA4 基因慢病毒载体构建与鉴定.

Objective To construct aud verify the RNA interlerence (RNAi) lentiviral vector of GATA4 gene. Methods We desigued two siRNA interlerence sequences of GATA4 gene aud connected them with pMGic7.1 vector after double enzyme digestion. The constructed vector was transfected into 293T cells, aud they were raudomly divided into the experimental mp, negative control up, aud blank control up. The expression of GATA4 was detected by Western blotting aud RT-qPCR, respectively Results The pMGic 7. 1 vector was correctly connected with one siRNA interlerence sequence, the gene sequence was as follows; CGAAACACCGGGGACATAATCACTGCGTAATCCTCGAGGATTACGCAGTGATTATGTCCTTTTTTGAATTCGGA (The bold part is the siRNA interference sequence), which met the requirements for the experiment. No target gene was added into the blank control group, and there was no protein band at 45 kDa. The relative expression levels of GATA4 protein in the experimental group and negative control group were 0. 739 0. 056 and 0. 997 0. 001, respectively, with statistically significant difference between the two groups (all P < 0. 05). The relative expression levels of GATA4 mRNA in the experimental group, negative control group, and blank control group were O. 362 0. 024, 0. 998 0. 001, and 0. 812 0.001, respectively; the relative expression of GATA4 in the experimental group was significantly lower than that of the negative control group and the blank control group (all P < 0. 05). Conclusion The GATA4 gene 1 entiviral vector is successfully constructed, and the GATA4 gene can be effectively inhibited in vitro. [ABSTRACT FROM AUTHOR]