Genetic Variation in the Calpastatin Gene and its Association with Growth Traits in Batur Sheep
Abstract
The present study aimed to investigate the association of the CAST genotype and growth traits in Batur sheep. Batur lambs were reared under an intensive feeding system. Bodyweight is measured monthly after weaning until six months of age. Blood representing thirty head were collected, genomic DNA was extracted as samples, and then 200 µl of whole blood samples were used. Specific primers were designed to amplify the CAST gene, samples were sequenced, then the researchers used the BioEdit program to identify any mutation. Calculation of genotypes, gene and allele frequencies, heterozygosities, and Chi-square test was performed. The analysis revealed a total of sixteen polymorphic sites in the CAST coding region. There are four alleles observed (A, G, C, and T), trans-versions at c.92T<G and c.285G>T loci, and transitions at c.214G>A, c.280G>A, c.301C<T, and c.371A>G. One individual disrupted the reading frame in the whole CAST sequenced. The genotype frequency analysis showed the highest predominance of the (TT, GG, CC, and AA) genotypes with frequencies (66.7 %, 62.9 %, 59.3 %, 66.7 %, and 55.6 %) as homozygous. In contrast, the heterozygous (TG, GA, GT, CT, and AG) genotypes were present at lower frequencies (29.6 %, 33.3 %, 37 %, 29.6 %, and 37 %), respectively with four difference alleles (T, G, C, and A). Post-weaning bodyweight till 6-month age of Batur sheep for the heterozygous genotypes at loci (c.92T<G, c.214G>A, c.280G>A, and c.301C<T) was slightly more massive than those carrying homozygous wild-type genotypes (c.285G>T and c.371A>G) with no significant differences (p > 0.5). The lambs with the heterozygous genotype had a significantly higher muscle percentage as a whole, compared to the other genotypes. Polymorphic sites were present in Batur sheep for the first time about the association with the CAST gene. However, the CAST gene might not be a useful marker for developing future selection programs in Batur sheep unless further investigation of the CAST gene and its interactions with other genes involving muscle growth and carcass traits are analyzed through association studies at a large scale.
Keywords: genetic variation, CAST gene, association, Batur sheep, growth traits
References
[1] Moradi MH, Nejati-Javaremi A, Moradi-Shahrbabak M, Dodds KG, McEwan JC. Genomic scan of selective sweeps in thin and fat tail sheep breeds for identifying of candidate regions associated with fat deposition. BMC Genet. 2012;13(1):10.
[2] Kemp CM, Sensky PL, Bardsley RG, Buttery PJ, Parr T. Tenderness—an enzymatic view. Meat Sci. 2010;84(2):248–56.
[3] Zhou H, Hickford JG, Gong H. Polymorphism of the ovine calpastatin gene. Mol Cell Probes. 2007;21(3):242–4.
[4] Wang KK, Yuen PW. Calpain inhibition: An overview of its therapeutic potential. Trends Pharmacol Sci. 1994;15(11):412–9.
[5] Huff Lonergan E, Zhang W, Lonergan SM. Biochemistry of postmortem muscle - lessons on mechanisms of meat tenderization. Meat Sci. 2010;86(1):184–95.
[6] Goll DE, Thompson VF, Li H, Wei W, Cong J. The calpain system. Physiol Rev. 2003 Jul;83(3):731–801.
[7] Temm-Grove CJ, Wert D, Thompson VF, Allen RE, Goll DE. Microinjection of calpastatin inhibits fusion in myoblasts. Exp Cell Res. 1999;247(1):293–303.
[8] Dedieu S, Mazères G, Poussard S, Brustis JJ, Cottin P. Myoblast migration is prevented by a calpain-dependent accumulation of MARCKS. Biol Cell. 2003;95(9):615–23.
[9] Koohmaraie M, Geesink GH. Contribution of postmortem muscle biochemistry to the delivery of consistent meat quality with particular focus on the calpain system. Meat Sci. 2006;74(1):34–43.
[10] Calvo JH, Iguácel LP, Kirinus JK, Serrano M, Ripoll G, Casasús I, et al. A new single nucleotide polymorphism in the calpastatin (CAST) gene associated with beef tenderness. Meat Sci. 2014;96(2):775–82.
[11] Byun SO, Zhou H, Forrest RH, Frampton CM, Hickford JG. Association of the ovine calpastatin gene with birth weight and growth rate to weaning. Anim Genet. 2008;39(5):572–3.
[12] [12] Chung H, Davis M. Effects of genetic variants for the calpastatin gene on calpastatin activity and meat tenderness in Hanwoo (Korean cattle). Meat Science. 2012;90(3):711-714. https://doi.org/10.1016/j.meatsci.2011.10.017.
[13] Greguła-Kania M, Gruszecki TM, Junkuszew A, Juszczuk-Kubiak E, Florek M. Association of CAST gene polymorphism with carcass value and meat quality in two synthetic lines of sheep. Meat Sci. 2019 Aug;154:69–74.
[14] Nassiry MR, Tahmoorespour M, Javadmanesh A, Soltani M, Far SF. Calpastatin polymorphism and its association with daily gain in Kurdi sheep. Iran J Biotechnol. 2006;4:188–92.
[15] Schenkel FS, Miller SP, Jiang Z, Mandell IB, Ye X, Li H, et al. Association of a single nucleotide polymorphism in the calpastatin gene with carcass and meat quality traits of beef cattle. J Anim Sci. 2006 Feb;84(2):291–9.
[16] Curi RA, Chardulo LA, Mason MC, Arrigoni MD, Silveira AC, de Oliveira HN. Effect of single nucleotide polymorphisms of CAPN1 and CAST genes on meat traits in Nellore beef cattle (Bos indicus) and in their crosses with Bos taurus. Anim Genet. 2009 Aug;40(4):456–62.
[17] Urbański P, Pierzchała M, Terman A, Kamyczek M, Różycki M, Roszczyk A, et al. The relationship between the polymorphism of the porcine CAST gene and productive traits in pigs. Can J Anim Sci. 2015;95(3):361–7.
[18] Gandolfi IG, Zambonelli P, Braglia S, Speroni C, Zappavigna A, Russo V, et al. Polymorphisms and haplotypes of calpastatin gene are associated with performance and carcass traits in Italian large white pigs. Livest Sci. 2012;147(1-3):77–83.
[19] Wang J, Elzo MA, Jia X, Chen S, Lai S. Calpastatin gene polymorphism is associated with rabbit meat quality traits. J Appl Anim Res. 2017;45(1):342–5.
[20] Zhou YG, Xiong Y, Yang CW, Jiang XS, Ran JS, Jin J, et al. 2017. Experimental verification of CAPN1 and CAST gene polymorphisms in different generations of Da-Heng broilers. BioMed Research International. 2017. https://doi.org/10.1155/2017/7968450.
[21] Johnston DJ, Graser HU. Estimated gene frequencies of GeneSTAR markers and their size of effects on meat tenderness, marbling, and feed efficiency in temperate and tropical beef cattle breeds across a range of production systems. J Anim Sci. 2010;88(6):1917–35.
[22] Bickerstafe R, Hickford JG, Gately K, Zhou H. Association of polymorphic variations in calpastatin with meat tenderness and yield of retail meat cuts in lambs. Proceedings of international congress of meat science; 2008; Helsinki, Finland.
[23] Prayitno. 2010. Analisis Genetik dan Kekerabatan Domba Batur dengan Domba Lokal dan Merino Menggunakan Marker RAP-DNA. Laporan Penelitian Hibah Doktor. Universitas Gadjah Mada, Yogyakarta.
[24] Nei M, Kumar S. Molecular evaluation and phylogenetics. New York: Oxford Univ Press; 2000.
[25] Vasconcellos LP, Talhari DT, Pereira AP, Coutinho LL, Regitano LC. Genetic characterization of Arberdeen Angus caĴ le using molecular markers. Genet Mol Biol. 2003;26(2):133–7.
[26] Noor RR. (2008). Genetika Ternak. Ed. Ke-2. Jakarta: Penyebar Swadaya.
[27] Enriquez-Valencia CE, Pereira GL, Malheiros JM, de Vasconcelos Silva JA, Albuquerque LG, de Oliveira HN, et al. Effect of the g.98535683A>G SNP in the CAST gene on meat traits of Nellore beef cattle (Bos indicus) and their crosses with Bos taurus. Meat Sci. 2017 Jan;123:64–6.
[28] Dagong MI, Cece S, Ronny RN, Rachmat H, Mohamad Y. Genetic polymorphisms of the coding region (exon 6) of Calpastatin in Indonesian sheep. Media Peternakan. 2011;190-195. https://doi.org/10.5398/medpet.2011.34.3.190
[29] Sumantri C, Diyono R, Farajallah A, Inounu I. Polymorphism of calpastatin gene and its effect on body weight of local sheep. JITV. 2008;13:117–26.
[30] brahim AH, Ismail IM, Shehata MF, El-Beltagy AR. Calpastatin Polymorphism in Barki Lambs and their Effects on Growth and Carcass Traits. J Am Sci. 2015;11(3) . Available from: http://www.jofamericanscience.org
[31] Armstrong E, Ciappesoni G, Iriarte W, Da Silva C, Macedo F, Navajas EA, et al. Novel genetic polymorphisms associated with carcass traits in grazing Texel sheep. Mesc. 2017. https://doi.org/10.1016/j.meatsci.2018.06.014.
[32] Dagong, M. I. A, Cece Sumantri, Ronny Rachman Noor, Rachmat Herman, Mohamad Yamin. (2016). Growth characteristics of Indonesian Thin Tail Sheep (TTS) based on Calpastatin (CAST) gene genotype variation. Advances in Environmental Biology. 2016;10(11):27-31.
[33] Kuryl J, Kapelanski W, Pierzchala M. Preliminary observations on the effect of calpastatin gene polymorphism in carcass trait in pigs. Anim Sci Pap Rep. 2003;2:87– 95.