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In this study, the complete mitochondrial genome of Marco Polo wild sheep was sequenced for the first time. It is 16 613 bp in length and possesses 22 tRNA genes, 13 typical mammalian protein-coding genes, two rRNA genes and one D-loop region. The whole genome's base composition is 33.7% A, 25.8% C, 13.1% G and 27.4% T, and the percentage of AT-rich regions is 61.1%.

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The CCNG1 gene encodes cyclin G1, which is an important cell cycle regulator and has been reported to be involved in reproductive biological processes, such as oocyte maturation and granule cell proliferation in mammals. But the study of CCNG1 in sheep has been rarely reported. To examine the effects of CCNG1 on estrous control and seasonal breeding in sheep, we first cloned and characterized the expression level of the sheep CCNG1 gene. Then by Real-time PCR, we detected and analyzed the expressions of CCNG1 gene at mRNA levels in the hypothalamus-pituitary-ovary (HPO) axis in different stages of an estrous cycle in Duo Lang sheep (non-seasonal breeding) and Merino sheep (seasonal breeding). The results showed that the open reading frame of the sheep CCNG1 gene is 885 bp in length and encodes 294 amino acids. Bioinformatic analysis indicated that the secondary structure of the sheep CCNG1 protein contained multiple phosphorylation sites and some Protein Kinase C phosphorylation sites. CCNG1 mRNA was identified in all tissues tested, with the levels in ovary and kidney higher than others. The expression profiles of CCNG1 in the HPO axis in different stages of an estrous cycle were similar in different sheep breeds: the expression levels of CCNG1 in the ovary, uterus, pineal gland and pituitary gland all peaked in the estrus phase. But there were significant differences for expression change extent of CCNG1 in ovaries in the oestrus and metestrus phase between different sheep breeds. The results suggested that CCNG1 probably participated in the regulation of estrous behavior and seasonal reproduction through controling the growth and development of follicles in sheep.

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FABP4 (Fatty acid binding protein 4) is a hot candidate gene in fat deposition and lipid metabolism and participates in the transport and metabolism of intracellular free fatty acids. We aim to study the role of FABP4 in fat deposition and metabolism of the rump fat in Altay sheep. In this study, bioinformatics method was used to analyze the protein sequence homology among 10 species, and RT-PCR was employed to detect FABP4 tissue profiling of Altay sheep. An animal model simulating the rump fat deposition and metabolism of Altay sheep was established by continuous starvation, and qPCR and iTRAQ (isobaric tags for relative and absolute quantitation) were used to detecte FABP4 mRNA and protein expression changes in the control and continuous starvation groups, respectively. Sequence analysis showed that FABP4 protein sequence is highly conserved among species, suggesting an important biological function during evolution for FABP4. The RT-PCR result confirmed that FABP4 mRNA was highly expressed in intestinal and rump fat, suggesting that FABP4 plays an important physiological role in fat tissues. We did not find significant differences in FABP4 mRNA and protein between control and continuous starvation groups (P>0.05), which indicates that FABP4 may not be the key gene in fat deposition and metabolism in Altay sheep.The results above lay a foundation for further studies of FABP4 in rump or tail fat.

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Studies have shown that clock gene Cry1 may have important roles in the endocrine process of seasonal reproduction in mammals. In this study, Duolang sheep (non-seasonal reproduction sheep breed) and Chinese Merino (seasonal reproduction sheep breed) were used to determine the expression change of Cry1 in hypothalamus-pituitary-ovary axis in different stage of estrous cycle by quantitative real-time PCR. The results showed that the Cry1 mRNA was expressed in all tested tissues, in which the expression levels in pineal gland and thyroid gland were higher than in other tissues. As far as different sheep breeds were concerned, the tissue expression profiles of Cry1 at different stage of estrous cycle were broadly similar. Besides hypothalamus, the expression levels of Cry1 in ovary, uterus, pineal gland, pituitary gland, and thyroid gland were all reached to peak in proestrus. The differences of expression change extent for Cry1 in vary, uterus, pineal gland, and pituitary gland in proestrus and oestrus were significant between different sheep breeds. The results suggested that Cry1 may play roles in switching on the estrus and seasonal reproduction.

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In order to analyze the correlation of tail fat deposition and two SNP loci on Ovis arise chromosome X and provide a theoretical basis for using molecular assisted selection technology in further low-fat sheep breeding, the breeds with extreme differences in tail types (Altay, Small Tail Han Sheep, Hu, Chinese Merino and Suffolk) were used to detect the polymorphisms of two SNP loci on X chromosome and the haplotypes with PCR-RFLP method. The results showed that the TT genotype at 59571364 locus and GG genotype at 59912586 locus were preponderant genotypes in thin-tailed Chinese Merino and Suffolk sheep flocks, while the percentage of the two genotypes in fat-tailed (fat-rmup) Altay and Hu flocks is less than 2%. Haplotype analysis showed that CA haplotype is the main haplotype, the percentage of CA is up to 55%, and the percentage of CA and TA haplotypes together was 88.33% in Altay sheep flock. These results suggest that there are great differences in the SNP distribution of the 59571364 and 59912586 loci among different tail-typed sheep flocks, which can be used as molecular markers in high or low fat sheep breeding.

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Fat tail or fat rump is one of essential traits for surviving in harsh environments, and the mechanism of fat deposition and its inheritable characters in sheep are still unclear. Therefore, the 59383635th locus on X chromosome in our unpublished chip data was chosen as candidate SNP, PCR-SSCP method was used to detect genotypes in five sheep breeds which have extreme differences in tail types (Altay, Small Tail Han Sheep, Hu, Chinese Merino and Suffolk), and the mathematical model was employed to analyze the correlation between the polymorphism and the trait of fat tail or fat rump. The results in this study showed that the high frequency of allele T exists in Altay flock, and the frequency of allele C appears to be particularly high in the thin tail sheep breeds. The result of mathematical model showed that the ratio of T/C increased exponentially with the increase of phenotype score. These results suggest that there is a big difference in the SNP distribution between fat tail (rump) and thin tail sheep populations, and the SNP can be used as an ideal molecular marker in high-fat or low-fat sheep breeding. However, the biological function of the SNP remains to be further studied.

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Integrin linked kinase (ILK) is a scaffold protein, which plays important roles in hair follicle development. The cDNA sequence of novel ILK gene in sheep was cloned by PCR method and analyzed by bioinformatics. Tissue expression profiling in eight tissues and temporal profiling at different wool follicle anagen stages in skin was analyzed. The results showed that the whole open reading frame (ORF) of ILK gene was 1 359 bp in length, which encoded 452 amino acids. Bioinformatic analysis indicated that the secondary structure of ILK gene was mainly made up of three ankyrin repeats and a kinase domain, and there were multiple phosphorylation and Protein Kinase C sites in this gene. The RT-PCR result confirmed that ILK mRNA was expressed in heart, liver, spleen, lung, skeletal muscle, skin, and small intestine, and the expression level was much higher in skin, spleen, and liver than others. The q-PCR analysis demonstrated that the ex-pression level of ILK was significantly increased from March to May (early follicle anagen initiation) in both sheep breeds, Chinese Merino and Kazakh sheep, and there were certain differences from June to October between the two breeds. The above results indicated that ILK gene may play key roles in regulating secondary follicle growth.

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In this study, PCR-SSCP analysis was used to identify genetic variation in IGFBP-3 gene in Chinese Merino and Kazakh sheep. A PCR product of 178 bp corresponding to partial intron1 illustrated three unique binding patterns by SSCP analysis. Frequencies of the genotype AA, AB, BB and allele A, B in Chinese Merino sheep were 0.70, 0.24, 0.06, and 0.82, 0.18 respectively , and they were 0.87, 0.13, 0.00, and 0.93, 0.07 respectively in Kazaka sheep. Sequence analysis revealed a G/T transversion at position 122 of the fragment. This polymorphic locus of IGFBP-3 gene was at Hardy-Weinberg dis-equilibrium (P<0.01) in the two breeds. Different genotypes slightly affected several wool traits of Chinese Merino sheep. The individuals of genotype AA, AB, and BB had no significant difference in post-shearing weight and clean wool rate. Sta-ple length (SL) was decreased with the genotype of AA, AB, and BB, and the difference between AA and AB was significant (P<0.01). Greasy fleece weight (GFW) and follicle density in individuals of genotype AA was significantly lower than that in individuals of genotype AB (P<0.01) and BB (P<0.05); Average fiber diameter (AFD) in individuals of genotype AA was significantly higher than that in individuals of genotype AB (P<0.01) and BB (P<0.05).

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The current study was designed to detect SNPs within BMP15 and BMPR-IB gene and investigate the effect of the genes on sheep litter size. Four sheep lines, HU-Yang, Chinese Merino monotocous, Chinese Merino multiparous for wool production and Chinese Merino multiparous for mutton production, were used in this study. Litter sizes were recorded for each ewe in the four lines. Primers for BMP15 and BMPR-IB gene were designed from database sheep sequence and polymorphisms were detected by PCR-RFLP method. The results showed that there was no polymorphism with BMP15 gene among the four lines, and there was an A / G SNP with BMPR-IB gene at base 746 among the four lines. Three types of genotype (BB, B+ and ++), based on A / G locus, were found within each line. The frequencies of genotypes were significantly different among the lines (P<0.001), with BB genotype primarily existing in HU-Yang, ++ genotype in Chinese Merino monotocous line, and B+ genotype in Chinnese Merino multiparous lines. The A / G mutation influence significantly the sheep litter sizes, and the BB and B+ ewes had significant higher litter sizes than ++ ewes. The results of present study showed simultaneously that the genotype of BMPR-IB was a perfect predictor of the sheep litter sizes. These results intensively indicated that BMPR-IB is a major gene to affect litter size in sheep, and could be used as the molecular genetic marker to select litter size in sheep.

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