生物信息学预测及人Elp3敲除序列表达载体的构建

Objective: Using bioinformatics methods to forecast and construct human Elp3 deletion plasmids and study its structure and expression level. Methods: Homology modeling was used to analysis human Elp3 protein sequences, and construct recombinant plasmids; double enzyme digestion and sequencing metheds was used to identify the plasmids. RT-PCR, western blot was used to detect the expression of Elp3. Results: We constructed four deletion recombinant plasmids and the Elp3-overexpression plasmid, which named PIRES2-EGFP-Elp3-d1, PIRES2-EGFP-Elp3-d2, PIRES2-EGFP-Elp3-d3, PIRES2-EGFP-Elp3-d4 and PIRES2-EGFP-Elp3. The results of double enzyme digestion and sequencing metheds are right, besides, we get the right result of mRNA: elp3-d1(1374bp), elp3-d2(1110bp), elp3-d3(1239bp), elp3-d4(636bp), elp3(1692bp) in RT-PCR and protein: Elp3-d1(50KDa), Elp3-d2(40KDa), Elp3-d3(45KDa), Elp3-d4(27KDa), Elp3(62KDa) in western blot. Conclusion: Human Elp3 deletion plasmids were constructed successfully.