Identification of calpastatin gene (CAST|AluI) polymorphisms in indonesian cattle breed using PCR-RFLP technique

Abstract

Calpastatin (CAST) is dependent proteases and a specific inhibitor of the μ-calpain and m-calpain function, were found in mammalian tissues. Calpastatin (CAST) gene is a gene that have functions in inhibiting muscle protein degradation and responsible for muscle hypertrophy. The purpose of this study is to identify polymorphism of the Calpastatin (CAST) gene in Indonesian local cattles (Bali, Madura, Pesisir, Aceh and Katingan). DNA polymorphisms within the calpastatin gene may lead to the phenotypic differences of cattle growth traits. A 624 bp of Indonesian local cattles calpastatin gene was successfully amplified using Polymerase Chain Reaction (PCR) technique. PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism) method will be used to identify polymorphism of calpastatin genes in local cattles. An AluI restriction enzyme will cut the PCR product into two different length fragments that are 474 bp and 324 bp and revealed two alleles system, G and C and three genotypes GG, GC and CC. All of Indonesian local cattles population are polymorphic in calpastatin gene. The highest GG genotype frequency was found in all Indonesian local cattles, but the CC genotype was not found in Aceh Cattle. The G allele frequency was found in Bali Cattle population (85,8%), Madura Cattle population (80,1%), Pesisir Cattle population (78,6%), Aceh Cattle population (80,8%), and Katingan Cattle population (85,0%). The allele C frequencies of Bali, Madura, Pesisir, Aceh and Katingan Cattle population are 14,2%, 19,9%, 21,4%, 19,2%, and 15,0%, respectively. The higest heterozygosity value was found in Pesisir cattle and Aceh cattle, while the lower heterozygosity value was found in Bali, Madura, and Katingan cattles population. Based on the analysis, Indonesian cattle breed (bali cattle, madura cattle, pesisir cattle aceh cattle, and katingan cattle) are polymorphic in calpastatin gene with three genotipes (GG, GC, CC) and two alelles (G and C).

Gen calpastatin merupakan salah satu gen yang berhubungan dengan keempukan daging yang berfungsi sebagai penghambat (inhibitor) enzim calpain. Identifikasi keragaman gen calapastatin (CAST|AluI) pada sapi lokal dengan menggunakan metode PCR RLFP masih terbatas. Tujuan penelitian ini adalah untuk mengetahui keragaman gen CAST|AluI pada sapi Bali, sapi Madura, sapi Pesisir, sapi Aceh dan sapi Katingan dengan metode Polymerase Chain Reaction - Restriction Fragment Length Polymorphism (PCR-RFLP). Sampel darah sapi yang digunakan sebanyak 282 sampel yang terdiri atas sapi Bali (102 sampel), sapi Madura (68 sampel), sapi Pesisir (49 sampel), sapi Katingan (50 sampel) dan sapi Aceh (14 sampel). Identifikasi keragaman gen CAST|AluI menggunakan pendekatan Polymerase Chain Reaction - Restriction Fragment Length Polymorphism (PCR-RFLP) dengan menggunakan enzim restriksi AluI. Data genotipe gen CAST|AluI dianalisis dengan pendekatan frekuensi genotipe dan alel serta pendugaan nilai heterozigositas.