Isolation and Characterization of Genes Involved before and after Puberty in Xiao Meishan Pig’s Hypothalamus
|Course||Animal Genetic Breeding and Reproduction|
|Keywords||Meishan pig Hypothalamic Puberty starts Differential display PCR Quantitative PCR Reverse Northern Molecular Cloning|
At present, domestic and foreign scholars and variety of animals puberty regulatory factor many studies to clarify the impact of steroid hormones, growth factors, neurotransmitters, body fat metabolism signal hypothalamic neurons release GnRH. But in addition to the above-described regulatory factor on porcine puberty start adjusting role, whether there are still unknown regulatory genes affecting pigs puberty start unclear. The purpose of this study by RT-PCR, differential display PCR (DDRT-PCR), reverse Northern (Reverse Northern), fluorescence quantitative PCR (real time PCR) and other molecular biology techniques, Meishan pig before and after puberty under hypothalamus differentially expressed unknown gene screening, identification, and their expression in different physiological and developmental stages variation linked, to explore the role of these genes in the pig puberty start mechanism, is rich in animal reproductive mechanism and early-maturing varieties selection election education provides a theoretical basis. The main findings are as follows: the test of birth, puberty, sexual maturity the purebred Xiaomeishan sow 3, to remove Health (0 days), puberty (70 days) (I group), as well as puberty (70 days old), sexual maturity (150 days of age) (II group) each three Meishan pig hypothalamus total RNA, from each the 10μl components of the total RNA pool 30 μl. Were composed of two groups of four RNA pools. Using 10 anchor fixed cited material, 100 kinds of primer combinations of 10 random primers material DDRT-PCR, a total of 12 differentially expressed strip with NCBI than on discovery which four sequences with known genes high degree of homology, respectively named as MS01 (355bp), MS02 (391bp), MS03 (241 bp), MS04 (281 bp). MS01 and porcine brain-specific vascular generation inhibitor (bpai) highly homologous homology of 98%, GenBank accession number for the the XM 0 01,925,295. MS02 pig brain fatty acid binding protein gene (fabp7) a high degree of homology, the homology of 93%, GenBank accession number DQ000160. MS03 and pig chromosome 13 section of unknown function sequence homology, the homology of 86%, GenBank accession number CU633348. MS04 with human collagen splice variants have strong homology, homology of 95%, and GenBank Accession No. DQ644568. Other sequences in the database become the same gene. Finally, by Reverse Northern, real time PCR qualitative and quantitative detection of gene expression level. The quantitative results show: Porcine Hypothalamus three developmental stages bpai mRNA expression levels showed a downward trend, and the gene expression levels in the newborn when compared with puberty and sexual maturity, the difference in expression was significantly (P lt; 0.05) The results suggest that the hypothalamus birth to sexual maturity in the development process, this factor is involved, pulsed release of hypothalamic neurons puberty nowadays provide access security, plays the role of the pipeline rapidly developing guide; fabp7 mRNA next hypothalamus three different developmental stages, the expression levels showed an upward trend, differentially expressed not statistically significant (P gt; 0.05), the experimental results suggest that fatty acid regulation factor in Meishan pig in its expression gradually increased with the growth of the age increasing incremented and accumulated in a process, this factor as a fatty acid receptor, and enhance the affinity of the fatty acid and its derivatives, to promote the uptake of fatty acids, suggesting that this factor can be further increased fat accumulation of pigs, while fat accumulated in further results directly affects pigs the nutritional status of the body, when the amount of fat accumulation reaches pigs start puberty desired body fat level, and then in collaboration with other regulatory factors participate pigs start of puberty.