Microsatellite Marker Of Cold-torlerance And Genetic Variation Of Candidate Gene In Guyuan Chicken

Guyuan chicken is the native populations spreading over cold region of northwestern of China. Cold - tolerance is a special character in Guyuan chicken population. In order to searching for linkage phenotype with the trait of cold tolerance, there are 766 individuals, including Tibetant chicken (cold-tolerance populations), Hesix chicken (non cold-torlerance populations), Wenchang chicken (hot-torlerance indigenous populations) and F2 population obtained from Guyuan hybrid with their parents (as the control population). The adoption of these populations stands for region-culture all in virtually cold and hot regions of China. There are 26 pairs of microsatellite primers have been employed to search for the linkage loci with cold tolerance. And the single nucleotide polymorphism and DNA sequencing were used to study on 3 candiadate genes: TYR gene, PPAR gene and NPY gene. Therefore, it could be great interest to know whether the differences in three genes of different chicken populations. So the aim of this study was to analyze the variation and polymorphism of the three genes, provide some important information to lipid metabolism regulation of cold tolerance, especially to accelerate the development of a cold tolerant strain of chicken in cold region of China, such as Guyuan and Tibet. The results were summarized as follows:(1) In crosspopulationsing experiment, Guyuan chicken had significance cold tolerance (P<0.01). There was probably cold-tolerance gene in Guyuan chicken and it would be inheritance. In control experiment, the feed intake of Hesix chicken was significant lower than that of Guyuan chicken (P<0.01). In Guyuan population, the egg yield of the black was significant lower than that of native Guyuan chicken (P<0.01). There were no significance differences in feed intake between them. The correlation coefficient of cold-tolerance and weight was 0.4235. But there were no significant difference in different individuals. So, there were no direct relation between weight and tolerance. There was no significance correlation between the cold tolerance and skin color, feather color (P>0.05).(2) Geneticstructure and diversity of five chicken populations were evaluated with 26 microsatellite loci. The result showed that there was abundant genetic diversity in three Chinese chicken populations. The average heterozygos value of Tibetant was the highest (0.822). Considerable population differentiation was observed and there was 45.5 % of the total genetic variation came from populations differences. The genetic distance of Guyuan chicken and Tibetant chicken was the most nearly. And they contained in one group. The Wenchang chicken and Hesix chicken contained to other group separately. Association analysis between the polymorphisms and body weight growth trait of Guyuan, at MCW169, MCW166, MCW351 and MCW073 locus, individuals with genotype BE, CD, BB and GG had higher body weight compared to individuals with other genotype, AD, DD and GK.(3) Two special microsatellite loci were found. At LEI212 locus, every individuals of Guyuan and Tibetant as cold torlerance populations had PCR products. But there were not products in Wenchang and Hesix populations. The chi-square of allelic analysis revealed that there was no significant difference between Guyuan and Wenchang populations (P>0.05). At MCW141 locus, the chi-square analysis revealed that there was no significant difference between Guyuan and Wenchang populations (P>0.05), but there was significant statistical difference between cold-tolerance and hot-tolerance populations (P<0.01). Aligned them with the sequence of GenBank, the two loci were located at Chromosomes 6 and 3, respectively. And the two loci were possibly related with cold-tolerance and hot-tolerance, which could be considered as the candidate loci and might be the effective loci for further study.(4) In this study, six loci of the gene exonl and one locus of 5' flanking region were analyzed by PCR-SSCP and DNA sequencing. The results indicated that there were polymorphisms only at TYR1 and TYR3 locus. At TYR1 locus, there was one silent mutation site: C to T, three genotypes (TT, CC and TC) was detected in three Chinese chicken populations respectively. Genotype CC wasn't detected in Hisex chicken. The chi-square analysis revealed that there was only significant difference between Wenchang and Tibent populations (P<0.01). At TYR3 locus, there was one mutation site: G to A, three genotypes (GG, AA and GA) was detected in Chinese local chicken populations and genotype AA wasn't detected in Hisex chicken populations. The chi-square analysis revealed that there was only significant difference between Wenchang and Guyuan populations (P<0.01). It was concluded that there was abundant variation of TYR gene in Chinese local chicken populations. It showed that there was no direct relation between cold tolerance and the variation of regions of exon1 and one locus of 5' flanking region of TYR gene.(5) In this study, the genetic variations of PPARA gene in 4 chicken populations (Guyuan, Wenchang, Tibetan and Hisex) were detected by PCR-SSCP and DNA sequencing. The results indicated that 6 different genotypes (namely, AA-EF) were detected in four chicken populations at P4 locus. Aligned them with the sequence AF163809, the nucleotide sequences of genotype AA, BB, AB and CC all revealed silent mutations in three Chinese populations. The nucleotide sequences of genotype DD and EF in Hisex showed several frame-shift mutations. It was implied that there were abundant variations of PPARαgene in chicken populations. Moreover, distribution of genotype frequency within PPARαgene was significant difference in 4 populations selected for special purpose in special region. It was implied that this locus could be probably utilized as an effective marker in marker assisted selection for layer, meat and egg concurrent and broiler populations.It showed that there was direct relation between cold tolerance and the variation of PPARA gene, but which needed to be confirmed further.(6) In this study, two loci of the gene exon2 and intron 2 of NPY (Neuropeptide Y) gene were analyzed by PCR-SSCP and DNA sequencing. The results indicated that there was significient polymorphism only at NPY2 locous. There was one mutation site: T to C at 1558 nt (Aligned them with GenBank accession No: 3JJM87298), which caused the 23th amino acid variation (Arg to Cys). And only a few genotypes BB were detected in four chicken populations. The chi-square analysis revealed that there was significant difference between cold tolerance populations and Wenchang populations (P<0.01). There was no significant difference between Hesix and other three populations (P>0.05). It showed that there was no direct relation between cold tolerance and the variation of NPY gene. Association analysis between the polymorphisms and body weight growth trait of Guyuan, at NPY2 locus, individuals with genotype AB had higher body weight compared to individuals with genotype AA.