pEGFP-C1-U6 plasmid vector-mediated expression of MDR1 shRNA Plasmid Construction.docVIP

 PAGE \* MERGEFORMAT 24 pEGFP-C1/U6 plasmid vector-mediated expression of MDR1 shRNA Plasmid Construction 【Abstract】 In order to construct pEGFP-C1/U6 vector-mediated MDR1 short hairpin RNA (short hairpin RNA, shRNA) expression plasmid for MDR1 fragment size of 19 bases, respectively, two pairs of oligonucleotides designed to form a double - chain after its turn connected with the U6 promoter of the pEGFP-C1 vector (named pEGFP-C1/U6), two pairs of DNA double-strand connected to form intermediate 9 base sequence intervals from the reverse complementary sequence to build Cheng can generate MDR1 short hairpin RNA plasmid. The results showed that: After digestion, the connection to build into a plasmid (pEGFP-C1/U6/A and pEGFP-C1/U6/B), confirmed by restriction enzyme digestion and sequencing to build a successful, without any mutation. Conclusion: The successfully constructed to express MDR1 shRNA plasmid vector pEGFP-C1/U6/A and pEGFP-C1/U6/B, the results of this study may have clinical reversal of multidrug resistance provides an effective method. Keywords: pEGFP-C1/U6 carrier Construction of pEGFP-C1/U6-Mediated Plasmid Expressing MDR1 shRNA Abstract To construct a plasmid expressing MDR1 short hairpin RNA (shRNA) mediated by pEGFP-C1/U6 vector, two coding sequences of 19 nucleotides were selected from MDR.Two pairs of oligonucleotides were designed for these two fragments.After annealing the formed double - stranded DNAs were ligated with plasmid pEGFP-C1/U6 (pEGFP-C1 vector with U6 promoter). The plasmids producing MDR1 shRNA were constructed from the inverted motif containing 9 spacers and four Ts. The results showed that the constructed plasmids were named pEGFP - C1/U6/A and pEGFP-C1/U6/B, and the constructs were identified by restriction and sequence analysis, no any base mutation was observed. It is concluded that plasmids of pEGFP-C1/U6/A and pEGFP-C1 / U6 / B expressing MDR1 shRNA were successfully constructed, providing a