| Rex rabbit was famous for its smooth, soft, keeping warm, non-fading and durable fur. Thus, it had long been very popular with consumers. The fur color was not only one of the most intuitive, heritable phenotypic characteristics, but also an important evidence to distinguish the animal species. The color formation in mammals was mainly controlled by heredity, and environment and feeding also played a role. Differentially expressed genes (DEGs) were achieved between chinchilla Rex rabbit & white Rex rabbit, and chinchilla Rex rabbit & black Rex rabbit by Solexa sequencing technology. The quantitative real-time PCR was used to validate the results of Solexa sequencing. PCR-SSCP and TA clone sequencing were used to analyse the selected DEGs, screened with rabbit fur color formation. The result provided some reference for further study of the genetic mechanism of Rex rabbit fur color. The main results were as follows:1. In this study, Solexa sequencing technology was used to analyze the gene expression of dorsal skin tissues collected from three full-sib Rex rabbits with different colors in order to explore the molecular genetics associated with fur color determination in domestic rabbits. Compared with the chinchilla (C) Rex rabbit control sample, the numbers of genes that exhibited greater than a 2-fold change in expression with a false discovery rate of ≤0.001 in the black (B) rabbit sample and white (W) rabbit sample were 1,809 and 460 respectively, and the number of common differentially expressed genes was 257. The quantitative real-time PCR showed gene expression trend of Tyrpl, Oca2, Dct, Slc24a5, Slc7al 1 and Bcl2 genes was consistent with that of Solexa sequencing data, indicating that the result of Solexa sequencing was reliable. Combined with bioinformatic analysis and related literatures, some genes were potentially related to fur color formation, such as Tyr, Tyrpl, Oca2, Dct, Slc24a5, Slc7all, bcl2 and so on. The alternative splicing and SNPs were also predicted in this study, which can be studied by further studies to investigate the relationship between them and the color formation.2. There were total 600 rabbits were used,180 of which were white Rex rabbits,420 were chinchilla Rex rabbits. Specific primers were designed to detect the polymorphism of 5 exons of Tyr gene used PCR-SSCP. The result showed that the exon 2 of Tyr in 62bp (the gene CDS 881bp) had an A>G mutation, and glutamate was changed to be glycine (GAG>GGG); the exon 3 of Tyr in 82bp (the gene CDS 1118bp) had an A>C mutation, and threonine was changed to be lysine (AAG>ACG). Bioinformatics were used to analyze the mutations of Rex rabbit Tyr gene exon 2 and exon 3 to compare the physicochemical properties of wild-type Tyr protein with mutant one. As a result, the transitions of base A in exon 2 didn’t alter the physicochemical properties and structure of TYR protein, and the transversion of base A in exon 3 only changed the transmebrane structure, but didn’t alter the physicochemical properties and structure of TYR protein. Although the two mutations changed the composition of TYR protein, the key amino acid sites of the structure and function of TYR protein were not affected.3. There were total 600 rabbits were used,180 of which were white Rex rabbits,420 were chinchilla Rex rabbits. Specific primers were designed to detect the polymorphism of 7 exons of Tyrpl gene used PCR-SSCP. The result showed that, the exon 2 of Tyrpl in 29bp (the gene CDS 483bp) had a C>T mutation, and the mutation didn’t change the amino acid. The TA cloning technology was used to analyze the 6 introns of Tyrpl gene of chinchilla Rex rabbit, and 53 mutations were found, whether these mutations will have an impact on the expression and function of Tyrpl gene, still need to be further studied. |
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