| The high incidence of gastrointestinal diseases in young rabbits has been restricting the development of domestic rabbit industry.At present,some domestic and foreign companies adopt methods of restricting feeding to increase the survival rate during the fattening period.However,the research on limited feeding methods is still insufficient,and the standards are implemented in production.The inconsistency often leads to problems such as poor feed restriction or hindered growth and development after excessive feeding.Therefore,this study explored the effects of different levels of feed restriction on the growth performance and immunity of meat rabbits,and whether the compensatory growth after 2 weeks of feed restriction can achieve fully compensatory growth,in order to seek a scientific restriction feeding mode.And using transcriptome sequencing technology to screen out genes that may be related to the development of skeletal muscle in meat rabbits,and analyze the expression characteristics of development-related regulatory factors in skeletal muscle of rabbits under different feeding levels,in order to explore the growth of skeletal muscle in meat rabbits under restricted feeding conditions.The developmental mechanism provides a theoretical basis.The main test results are as follows:1.In the experiment,198 35-day-old meat rabbits were randomly divided into 3 groups,66 in each group,each half male and female,namely the control group,the 15%restricted feeding group Ⅰ and the 30%restricted feeding group Ⅱ.A feeding experiment of 3 weeks of restricted feeding and 2 weeks of compensatory growth was carried out.The results showed that there was no significant difference in the weight of the meat rabbits in the restricted feeding group Ⅰ compared with the control group(P>0.05),while the weight of the restricted feeding group Ⅱ showed a significant decrease(P<0.05)and there was no complete compensatory growth after 2 weeks of compensation feeding;during the restricted feeding period,the feed-weight ratio of the two groups of restricted feeding decreased significantly(P<0.05),and the health risk index of the restricted feeding group Ⅰ was 9.09%,which was significantly lower than that of the control group(P<0.05);after the end of feed restriction period,the jejunal chymotrypsin,alkaline phosphatase activity and sIgA level of feed restriction group Ⅰ increased significantly,while the sIgA and IgG levels of restricted feeding group Ⅱ decreased significantly(P<0.05),and there was no significant difference compared with the control group after the end of the compensation period(P>0.05);feed restriction group Ⅰ significantly increased the level of ileal anti-inflammatory factor IL-10(P<0.05),while after the end of the compensation period,the level of pro-inflammatory factors IFN-y and TNF-α increased significantly in group Ⅱ(P<0.05);the two groups of restricted feeding can significantly reduce the MDA content in the jejunum in each phase of the experiment.The levels of T-AOC and T-SOD in the restricted feeding group Ⅱ decreased after the restricted feeding period(P<0.05),while the T-SOD level increased significantly during the compensation period(P<0.05).Therefore,the 15%feeding level will not have a significant impact on the final body weight of the meat rabbit,on the contrary,it can significantly reduce the diarrhea rate and mortality of the meat rabbits,and improve the digestion and antioxidant capacity of the meat rabbits to a certain extent,while the 30%feeding level makes The development of skeletal muscle is blocked.After 2 weeks of compensatory growth,complete compensatory growth cannot be obtained.The protein synthesis of sIgA and IgG is blocked during the feed restriction period.After compensation,inflammatory factors increase due to overeating stress.2.Further analyze the effect of feed restriction on the growth and development of skeletal muscle,and use HE staining to measure and count the cross-sectional area of muscle fibers.In addition,real-time fluorescence quantification and Wes technology were used to detect the mRNA and protein expression of key factors in the PI3K/Akt signaling pathway.The results showed that the final body weight and muscle fiber area of the restricted feeding group Ⅱ were significantly reduced(P<0.05),and the expression of IGF1 gene in the restricted feeding group Ⅱ was found to be significantly reduced(P<0.05),while the expression of IGF2 was not significant among the groups(P>0.05).There was no change in the MSTN and FOXO1 gene levels in the feed restriction group Ⅰ(P>0.05),while the MSTN and FOXO1 mRNA expression levels in the feed restriction group Ⅱ were significantly increased(P<0.01)and significantly increased(P<0.05).The protein content of p-Akt and p-p70 in the restricted feeding group Ⅱ decreased significantly,while the p-Akt and p-p70 levels in the restricted feeding group Ⅰ did not change significantly(P>0.05).It shows that 15%feed restriction did not significantly affect the body weight and skeletal muscle development of meat rabbits,while 30%feed restriction level affects the growth and development of skeletal muscle of growing meat rabbits.This process may be achieved by regulating the PI3K/Akt signaling pathway.3.Through transcriptome sequencing analysis,the differential genes were screened with log2fold change|≥1&P<0.05,and it was found that there were 615 DEGs in the skeletal muscle tissues of meat rabbits in the control group and the restricted feeding group Ⅱ,of which 425 were up-regulated genes and 190 were down-regulated genes.Through the GO and KEGG functional enrichment analysis,it is found that there are 28 differential genes related to muscle development in GO enrichment,including IGFBP5,BARX2,MYH8,LMCD1,TNNI1 and other genes.KEGG analysis found that the main enriched signaling pathways of differentially expressed genes related to muscle development include PI3K/Akt signaling pathway,MAPK signaling pathway,and Hedgehog signaling pathway.Differential gene metabolism pathways include propionic acid metabolism and fatty acid degradation metabolism,including MYC,KIT,TBCID1,PAKAG3 and other genes.After a comprehensive analysis,it was found that genes such as MYH8,LMCD1,TNNI1,TBC1D1,PRKAG3 may be related to the growth and development of skeletal muscles in meat rabbits under restricted feeding conditions.4.The full length of TNNI1,a key gene for skeletal muscle development,was further cloned.Bioinformatics analysis revealed that TNNI1 is an unstable hydrophilic protein with 15 potential phosphorylation sites.The muscle tissues of 3-day-old young rabbits were collected,digested with collagenase,and purified by differential adhesion method to obtain myoblasts.The cells were obviously spindle-shaped and in good growth state.RT-PCR and immunofluorescence techniques were used to identify the markers MyoD,Myf5 genes and Desmin was positively expressed.The TNNI1 gene overexpression and interference experiment was further carried out,and it was found that after TNNI1 overexpression,the skeletal muscle development-related genes ANKRD2,CSRP3,MYOZ2 and BARX2 gene expression increased significantly(P<0.05).After TNNI1 interference,the skeletal muscle development-related genes ANKRD2,CSRP3,MYOZ2 and BARX2 gene expression decreased significantly(P<0.05).The results showed that changes in the expression level of TNNI1 can cause corresponding changes in the expression of muscle development-related genes,which may play a role in the process of nutritional regulation of muscle development in meat rabbits.In summary,15%feed restriction can reduce the rate of diarrhea and mortality on the basis of ensuring the growth performance of meat rabbits,and improve the digestion and antioxidant capacity of meat rabbits to a certain extent.However,the 30%feed restriction level hinders skeletal muscle development may be achieved by regulating the PI3K/Akt signaling pathway.At the same time,transcriptome sequencing screened some candidate genes related to skeletal muscle growth and development.The preliminary analysis results of TNNI1 gene indicated that it may play a certain role in the process of nutrition-regulating muscle development of meat rabbits. |
