Construction of a cellular model for detecting utrophin expression

摘要

Duchenne muscular dystrophy is caused by the absence of the muscle cytoskeletal protein dystrophin. Utrophin is an autosomal homologue of dystrophin, and overexpression of utrophin is expected to compensate for the dystrophin deficit. Our aim is to establish a convenient method for observation the expression of utrophin and a cellular model for drug screening of duchenne muscular dystrophy. Method: Obtain three fragment of utrophin gene at exon 18 and exon 19 and the sequence surround them by PCR. TETC gene (sequence contained tetracysteine motif) was inserted in 3 ' of fragment I. The target gene was constructed with pGPKneobpA-LoxP-A and fragments I, II, III, and was transformed into C2C12 cells by electroporation. To identify homologous recombinants, transfected C2C12 cells were screened by G418, FLASH staining and PCR detection. Result: Two homologous recombinated clones of C2C12 cells were obtained. Conclution: we have constructed a cellular model for detecting utrophin expression.