Study On Cloning Of IGFBP Family Genes And Tissue Expression Pattern In Nanjiang Yellow Goat

As a complex growth regulator shaft between IGFs and IGFBPs, IGFBPs regulate the IGFs biological activity by binding to IGFs with different affinity to achieve the regulation of animal growth and development. In this study, using Nanjiang yellow goat as experimental material and collected liver, heart, lung, longissimus dorsi muscle, triceps brachii muscle and semimembranosus muscle at five different postnatal periods(3,30,60,90and120days after birth, two males and two females at3day and three males and three females at other ages). The CDS of goat IGFBP-2、IGFBPS-3、 IGFBP-4and IGFBP-6genes were cloned from liver tissue. The different expression pattern of IGFBP-2, IGFBP-3, IGFBP-4and IGFBP-6genes in genders, tissues and ages were identified by real-time fluorescence quantitative PCR method. The different expression pattern of IGFBP-2protein in liver at different stages and in different tissues at120day were detected by western blotting technology. The main results were showed as follows:(1) The sequence lengths of IGFBP-2(JQ341160), IGFBP-3(JQ341161), IGFBP-4(JQ341162) and IGFFBP-6(JQ341163) were954bp,882bp,777bp and711bp, respectively, and encoded317,293,258and272amino acid residues correspondingly. The CDS sequences and the amino acid sequences of goat IGFBP-2、IGFBP-3、 IGFBP-4and IGFBP-6genes shared more than91%homology with those of sheep and cattle, and shared68%～94%homology with those of other species.(2) The deduced amino acid sequence of IGFBP-2、IGFBP-3、IGFBP-4and IGFBP-6contained hydrophilic region and signal peptide sites, but no transmembrane domains, excluding IGFBP-4which had one, also all four protein sequences had insulin growth factor-binding protein homologues domain and thyroglobulin type I repeats, and their main post-translational modifications were phosphorylation and glycosylation. Amongst the three kinds of secondary structure in goat IGFBP-2、IGFBP-3、IGFBP-4and IGFBP-6protein, the proportion of random coil was the highest(68.14%,72.70%,68.99%and78.39%), followed by alpha helix(18.30%,15.02%,21.32%and17.37%), and the extended strand(13.56%,12.29%,9.69%and4.24%). Additionally, phylogenetic analyses indicated that the goat IGFBP-2、IGFBP-3、IGFBP-4and IGFBP-6protein had close evolutionary relationships with sheep and cattle.(3) The expression levels of the four genes were detected in liver, heart, lung, longissimus dorsi muscle, triceps brachii muscle and semimembranosus muscle tissues at different stages, and existed significant differences between the relative expression levels. The relative expression in the liver and longissimus dorsi muscle were higher than in the other tissues, and showed a similar developmental regular pattern in the different tissues and different growth stages.(4) The western blotting analysis of IGFBP-2protein indicated that the relative content showed an upward trend in the liver at different stages, and it was higher in the liver and longissimus dorsi muscle than in other tissues at120day. These results were in consistent with the mRNA expression level.