Cloning the partial cDNAs of μ-calpain and m-calpain from porcine skeletal muscle

Abstract

Calpains are non-lysosomal proteases involved in myofibrillar protein degradation. To facilitate studying the expression of the porcine calpain genes and their influence on protein accretion, we have cloned partial cDNAs for μ- and m-calpain from porcine skeletal muscle via PCR amplification. A 289 bp fragment for μ-calpain and a 629 bp fragment for m-calpain were cloned into the EcoRV site of pBluescript II KS+ vector. The nucleotide sequence for porcine μ-calpain and m-calpain were 92% and 90% identical to corresponding regions of rabbit μ- and m-calpain, respectively. The deduced amino acid sequences for both μ- and m-calpain share 94% identity with respective rabbit μ- and m-calpains. Isoform specificity was validated by Southern hybridization of μ- and m-calpain probes with cloned μ- and m-calpain fragments and Northern hybridization with pig muscle mRNA. These clones will be used to evaluate the role of calpain expression in muscle hypertrophy.