| Animal growth and development are complex and sophisticated biological metabolic processes, in which genetic genes and hormones make an important role. In the growth control system, the regulation of various factors on the growth is mainly achieved through the growth hormone axis. The SNPs and mRNA expression of some key genes of growth hormone axis were determined in the present study, with attempt to verify the relationship between these key genes and Guangxi Bama mini-pig dwarfism. The results of the experiment were showed as follows:1:The serum GHR and IGF-Ⅰlevels on day1,30,180of age in Bama mini-pig and Landrace were measured by Elisa, respectively. The results showed that serum GHR and IGF-Ⅰlevels in Landrace were significantly higher than those in Guangxi Bama-mini pig. The serum levels of GHR on day1,30,180of age in Landrace were7.94,4.82and1.89times higher than those in Guangxi Bama mini-pig, respectively. Meanwhile, the serum levels of IGF-Ⅰon day1,30,180of age in Landrace were2.91,1.73and1.24times higher than those in Guangxi Bama mini-pig, respectively. 2:High-resolution melting (HRM) method was used to analyze the SS, GH, JAK2, IGF-I gene coding sequence polymorphisms (SNPs) of the Bama mini-pig and Landrace genomics as a reference to Huanjiang Xiang pig. The results showed that:(1) The SS, GH, JAK2and IGF-I gene in Guangxi Bama-mini pig were gene singlet IGF-I, SS, GH exon2and JAK2exon14,17in Landrace were polymorphisms.(2) Compared with Landrace, the differeces of genotype frequencies on two loci of SS gene and one locus of GH gene in Guangxi Bama-mini pig were significant (P<0.01).There were9different loci of JAK2gene in2breeds, and the genotype frequencies in8of those9loci reached the significant level.(3) The whole CDS of SS, JAK2, IGF-I and partical sequence of GH without exon4were obtained by splicing each exon together from HRM. Sequence alignment shows that the mutations in SS, GH, IGF-I gene were all synonymous mutations, only the615bp/1874bp/3364bp loci in JAK2were missense mutations.3:The cloned Bama-mini pig GHR cDNA contained1917bp, encoding638amino acid residues,which had a homology rate of99.6%compared with Landrace GHR cDNA sequence. The amino acid sequence alignment results showed that the coding region in two breeds is inconsistent in total of amino acids on the six sites. Of which, I309V was located in the tyrosine kinase binding domain, C390Y, S527N, and D561N were in GHBP binding region, the rest two K205R, A217V were in the extracellular domain. The predicted secondary structures of the growth hormone receptor showed that the K205R, A217V mutations did not make much impact on the protein secondary structure of the ligand binding domain. However, the models alignment showed the72～79amino acid peptide had separated. The GHR gene cloned from Guangxi Bama mini-pig and the Landrace were successfully ligated to pEGFP-C1vector, respectively to constructed eukaryotic expression vectors which will be used to study the effects of mutations in GHR gene on the down-stream regulatory process.4:Real-time PCR was used to analyze the expression situation of GHR, JAK2and IGF-I on day1,30,180of age in Guangxi Bama-mini pig and Landrace. The results showed that the expression level of3genes in Bama-mini pigs was reduced from day1to30of age, and which was increased in Landrace. The expression levels of the3genes on day1,30,180of age were significant higher in Landrace than in Bama mini-pigs (p<0.05). The significant difference of GHR, JAK2, and IGF-I gene expression levels on day1of age between this two breeds indicated that different expressions of these genes might occur before birth.In conclusion, it is speculated that GHR, JAK2gene is closely related to the dwarfism of Bama mini-pig. The hormone levels of growth axis, especially the reduction of GHR, JAK2and IGF-I secretion in the early growth stage and the down-regulated expression of GHR and JAK2may affect together on dwarfism of Guangxi Bama mini-pig. |
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