Transcriptomics Analysis Of Skin From XJ And SH By Analyzing Qualitative Character Of Wool

In this study, we used Xinji fine wool sheep and small tail han sheep as the research object to clarify the extreme difference of wool traits between these two varieties by detecting their phenotypic traits, skin histology and wool growth relative hormone serolog. Using transcriptome sequencing techniques to sequencing their skin tissues, then, assembling the measured sequence to compared with the reference genome and transcripts. Screening two differentially expressed genes from two sheep skin tissues, annotating them by GO and KEGG to get the biological function enrichment and specific metabolic pathways of differentially expressed genes. By studying on variable sites and differential splicing to dig out potential mutation site, mutant gene and variable spliceosome that related to wool traits difference, then choosing the keratin associated protein directly connect with wool traits to analysis the quality characters and genetic diversity, providing a theoretical basis for wool traits with molecular breeding of wool traits. The results are as follows:(1) Measured wool traits, histology and hormone in blood of Xinji fine wool sheep(n=6) and small tail han sheep(n=6). The results showed Xinji fine wool sheep’s fineness was significantly thinner than small tail han sheep, difference was extremely significant (P<0.01), the length of small tail han sheep’s wool was longer than Xinji fine wool sheep, difference was significant ((P<0.05). In the growth process of wool, the growth rate of small tail han sheep was faster than Xinji fine wool sheep. In terms of determination of hormones, two varieties of sheep’s growth hormone (GH) content is low, insulin-like growth factor (IGF-1), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), fibroblast growth factor (FGF-5), melatonin (MT), growth hormone (GH) and thyroid hormone (T3) were changed in different months, and there is a big difference. In the follicle density, compared these two sheep in the same month and same position, Xinji fine wool sheep was significantly bigger than small tail han sheep. Small tail han sheep was higher than Xinji fine wool sheep in skin thickness (P<0.01)(2) Selected the Xinji fine wool sheep(n=3)and Small tail han sheep(n=3)by high fulx sequencing of skin tissue, after filtration of sequencing data, we can get the high quality sequencing of lOObp of Xinji fine wool sheep is 0.6Gb, while Small tail han sheep is 0.56Gb. The inter comparison rate of sequencing and reference gene is 87.57% and 87.68%.Between two transcriptome libraries found 710 differentially expressed genes; 356 differentially expressed genes is up-regulated and 354 genes were down-regulated. By GO annotation of differentially expressed genes,416 differentially expressed genes commented to 208 Got term and formed 29 secondary item.118 genes were annotated in cell component,199 genes were in biological process and 321 genes in molecular function. By metabolism pathway annotation of differentially expressed genes,230 genes were annotated to KEGG database and gained 193 metabolic pathways,5 of them shown significant enrichment.(3)122,944 SNP sites and 5,683 Indel sites were detecting from Xinji fine wool sheep.129,074 SNP sites and 6,153 Indel sites were detecting from Small tail han sheep. During the process of variation function annotation, function annotation of Xinji fine wool sheep were 91,797 SNP sites and 5,684 Indel sites. Function annotation of Small tail han sheep were 129,091 SNP sites and 6150 Indel sites. Detected the potential functional annotation of objective traits related to variable sites between these two species by the threshold (Threshold) p-value<0.05, screened difference 5,236 SNP sites and 257 Indel sites. Detected the objective traits related to variable sites that located in mutant gene’s annotation and screened 155 mutated genes associate with protein changes. By GO annotation, gained 71 GO terms and 9 of them shown differentially expressed genes significant enrichment on mutation differentially expressed genes. By metabolism pathway annotation of mutation differentially expressed genes,40 genes were annotated to KEGG database and gained 88 metabolic pathways,1 of them shows significant enrichment. Detected the objective traits related to alternative splicing gained 76 differentially expressed genes, 93 spliceosome transcripts.(4)Analyzed of genetic variation of genen of keratin-associated proteins from Xinji fine wool sheep, we can find gene KAP24.1 T�'C mutation occurred in cDNA348bp, C�'T mutation occurred in cDNA414bp, both of them belong to missions mutation. Respectively Proline (Pro) was mutated to Leucine (Leu); Serine (Ser) to Proline (Pro); gene KAP13.1 T�'C mutation occurred in cDNA291bp, C�'T mutation occurred in cDNA469,528bp, did not cause Amino Acid Sequence change, belong to different mutation. In population genetics analysis, these two target gene sites keep in hardy-weinberg balance, and both in medium polymorphism(P<0.5); In gene KAP24.1,BB genotype has a significant effect on amount of wool production and extension length(P<0.05);In T291C site of gene KAP13.1.CC genotype has a significant effect on extension length, in C469、528T site of gene KAP13.1 NN genotype has a significant effect on amount of wool production and extension length(P<0.05).In summary, this study illustrates the difference of phenotypic traits, skin histology and wool growth relative hormone serolog between Xinji fine wool sheep and Small tail Han sheep.Selected differentially expressed genes and mutate genes by high fulx sequencing of skin tissue transcriptome to provide a theoretical data for further study of the growth and quality of wool. Meanwhile, analyzed genetic variation of Xinji-fine-wool sheep’s keratin associated protein to lay a theoretical basis for further study of molecular breeding of Xinji fine wool sheep.