Construction and identification of the eukaryotic expression plasmid for MeCP2 specific shRNA.

Objective To construct and identify the eukaryotic expression plasmid psilencer-MeCP2. Methods According to the Homo sapiens methy1-CpG binding protein 2(MeCP2) mRNA sequence, four strands of oligonucletide were designed and chemically syntheized, then annealed into two double strands in vitro. The two DNA strands were ligated with plasmid vector respectively, named psilencer-MeCP2/Ⅰ and psilencer-MeCP2/Ⅱ. These plasmaids were transformed into compenent cells(E.coli JM 109). After selective culture, recombined plasmids were extracted from E.coli, sequence was used to make sure that the two strands were inserted into plasmaids in correct sequence and direction respectively. Results The synthesized strands of oligonucletide contained correct and complete sequence of the shRNA, the MeCP2 specific shRNA has been inserted into eukaryotic expression. Conclusion The eukaryotic expression plasmid psilencer-MeCP2 has been constructed successfully.