Cloning, characterization and expression of myostatin (growth differentiating factor-8) gene in broiler and layer chicken (Gallus gallus)

Abstract

A study was conducted to characterize myostatin gene in broiler and layer chicken and to explore mRNA expression profile in these two varieties of chicken. The myostatin cDNAs of broiler and layer varieties were cloned and sequenced. The total length of the cDNA was 1128 bp. The differences of nucleotides between PB-1 broiler and IWI layer were C > 65 > T, C > 306 > T and C > 1094 > T while those between CB broiler and IWI layer were C > 65 > T, C > 195 > G, G > 234 > A, C > 306 > T, T > 939 > C, C > 961 > T, G > 966T and C > 1094 > T. The amino acid differences of myostatin protein between PB-1 and IWI strains were alanine > 22 > valine and proline > 365 > leucine while those between CB and IWI strains were alanine > 22 > valine, histidine > 321 > tyrosine and proline > 365 > leucine. The phylogenetic study revealed closeness of PB-1 and control broiler forming a cluster, which was also closely related to IWI layer chicken formed a separate cluster. The gene was cloned and expressed in E. coli. The gene expression profile in muscle was different between broiler and layer strains. Between two broiler strains, the pattern of expression was similar. Between IWI layer and either PB-1 or CB broilers, differences in expression was found at different time points, particularly at second, fourth and seventh weeks of age. The myostatin expression was significantly associated with body weights in chicken. It is concluded that myostatin gene sequences varied at nucleotide as well as amino acid level between broiler and layer chicken varieties and the gene also expressed differentially in these two varieties.

Comparative analysis of silencing expression of myostatin (MSTN) and its two receptors (ACVR2A and ACVR2B) genes affecting growth traits in knock down chicken

“…The GAPDH was used as reference gene. The amplification and comparative analysis of mRNA expression was performed by calculating 40-Δct in both knock down and control groups according to the method mentioned in the literature 45 , where, ∆ C t = Average C t of target gene – Average C t of reference gene.…”

“…The protein samples were electrophoresed in Tris-Glysin-SDS buffer (pH 8.3) at 60 V in 12% polyacrylamide gels of which one set of gels were stained with coomassie brilliant blue for visualization of resolved protein bands and another set of gels were used to transfer proteins onto PVDF membrane in the semidry transfer apparatus (Biorad) at 100 V for 1 hour. Then, the membrane was treated with anti rat IgG-HRP conjugate (Sigma) diluted to 1:1000 in TBS Tween buffer and visualized by treatment with DAB substrate using standard protocol 45 .…”

Myostatin mRNA expression and its association with body weight and carcass traits in Yunnan Wuding chicken

“…However, breed plays an important role in determining carcass characteristics and muscle mass. Breed significantly affects BW (Shahin and Elazeem, 2005;Jaturasitha et al, 2008;Bhattacharya et al, 2015), carcass weight (Ojedapo et al, 2008;Olawumi and Fagbuaro, 2011), breast and leg muscle weight fat and edible giblet weight (Ojedapo et al, 2008), and back and drumstick weights (Ojedapo et al, 2008) in chickens. Consistent with those reports, CW, BMW, LMW, AFW, DP, and EP were significantly lower in Wuding chickens than in broilers.…”

Production of Transgenic Chimeric Chicken from Cryopreserved Primordial Germ Cells and its Validation by Developing shRNA Transgenic Chicken Chimera

“…The GAPDH gene was used as internal control for which one pairs of primers, namely, QGAPDHF: 5'-CTGCCGTCCTCTCTGGC-3' and QGAPDHR: 5'-GACAGTGCCCTTGAAGTGT-3' for GAPDH gene was designed from the chicken cDNA sequences of the GAPDH gene (Accession No. AF047874) with DNASTAR software to co-amplify a 119 bp GAPDH fragment 34 . The surface markers of PGCs such as Nanog, SOX2, cPouV, Sox2, cMyc, Klf4 and cvh, BLIMP1, Myc and Klf4 were analysed for characterization of PGCs [46][47][48] (Table 6).…”