| Non-specific digestive disorder is an important disease in rabbit breeding, that is no specific drugs or prevention effects due to unknown etiology. The development of modern biotechnology provides a new way to explore the pathogenesis and prevention. IL-1β and TNF are important pro-inflammatory cytokines that play key biological roles in the induction and regulation of inflammatory responses. In the present study, the SNPs of IL-1β and TNF genes were firstly detected by directing sequencing of all exons, and then the candidate SNPs were genotyped by HRM. Association analysis between SNPs of IL-1β and TNF genes and non-specific digestive disorder was conducted by Case-Control study; using fibre-deficient diet ration to induce non-specific digestive disorder in New Zealand Rabbits, IL-1β and TNF genes mRNA relative expression in colon, ileum and sacculus rotundus tissues were detected, and the relationship of mRNA relative expression in different severity groups, different genotypes and different tissues were analysed; association analysis between the candidate SNP of TNF gene and day weights was also conducted. The experiment results were as follows:(1) Two SNPs located in exon4were detected in IL-1B gene, c.202G>A and c.223C>A, respectively;8SNPs were detected in TNF gene,2SNPs located in exon1(c.56A>G and c.87C>T),3SNPs located in intron1and3SNPs located in intron2. According to the SMART, SignalIP4.1and PANTHER online software and△RSCU value ananlysis, IL-1B c.202G>A, TNF c.56A>G and c.87C＞T were finally selected to study.(2) Allele A (OR=4.817, P=0.022) and genotype GA (OR=5.000,P=0.020) of IL-1B c.202G>A could increase the risk of developing non-specific digestive disorder in New Zealand Rabbits; TNF allele G of c.56A>G (OR=2.208, P=0.018), allele C (OR=2.005, P=0.0001) of c.87C＞T and genotype CC (OR=2.823,P=0.002) of c.87C＞T could increase the risk of developing non-specific digestive disorder in New Zealand Rabbits; moreover, haplotype H1(AT) and diplotype H1H1(ATAT) of TNF could decrease the risk of developing non-specific digestive disorder in New Zealand Rabbits.(3) Along with the increasing severity of non-specific digestive disorder,1L-1B and TNF genes mRNA relative expression was gradually increased in three tissues, and in colon the pairwise difference reached statistical significance between severity group and health group (P<0.05). These showed that IL-1B and TNF genes mRNA relative expression were closely related to the severity of non-specific digestive disorder.(4) The different genotype’s mRNA relative expression of IL-1B c.202G>A was not reached statistical significance in three tissues (P>0.05), that showed there was not significant association between polymorphism of IL-1B c.202G>A and IL-1B mRNA relative expression; genotype AG of TNF c.56A>G had the higher mRNA relative expression than genotype AA in colon and illume, and the pairwise difference reached extremely significance in illume (P=0.003); the different genotypes mRNA relative expression of TNF c.87C＞T was not reached statistical significance in three tissues (P>0.05); but TNF H2H4(ACGC) diplotype had significantly higher TNF mRNA relative expression than other three diplotypes in colon and illume (P<0.05), these showed that there was association between diplotypes and TNF mRNA relative expression undering the combined effect of TNF c.56A>G and c.87C＞T.(5) IL-1B and TNF genes mRNA relative expression in sacculus rotundus was extremely significantly higher than colon and illume (P<0.01), in colon was the lowest, and the difference between colon and illume was not significant (P>0.05). These showed that IL-1B and TNF genes mRNA relative expression were different in three tissues. Moreover, correlation analysis revealed that there was positive correlation between IL-1B and TNF genes mRNA relative expression (r>0, P＜0.05).(6) The different genotype’s day weight of TNF c.87C＞T was not reached statistical significance (P>0.05), this showed that there was not significant association between genotypes of TNF c.87C＞T and day weights. The present study revealed that the locus of IL-1B c.202G>A and TNF c.56A>G, c.87C>T should be considered as important candidate genetic locus associated with non-specific digestive disorder. |
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