Cloning of the human β-globin MAR and construction of the pCAT-MAR expression vector / 新乡医学院学报

ABSTRACT

Objective To clone the human β-globin matrix attachment region(MAR) and construct the mammalian animal expression vector pCAT-MAR, which contains the MAR and CAT reporter gene.Methods The human genomic DNA was extracted through phenol/chloroform and precipitated with ethanol, followed the MAR was amplified through PCR using the primers designed according to the GenBank sequence. After identified by agarose gel electrophoresis, sequenced and analyzed by the software, the PCR products were cut with restriction enzymes and ligated into the pCAT3-control vector to construct the pCAT-MAR vector. Results About 770bp band appeared in the agarose gel electrophoresis, the similarly compared with the published MAR sequence was 99.9%, the DNA fragment has the MAR typical features. The pCAT-MAR vector was demonstrated to be right through digestion with the corresponding enzymes and agarose gel electrophoresis. Conclusion The humanβ-globin MAR is cloned through PCR, and the expression vector pCAT-MAR containing the MAR is successfully constructed.