Analysis Of The Effect Of Combined Probiotic Complex And Essential Oil Additive On The Growth Of Hu Sheep By RNA-seq

Hu sheep,as an important protection breed of livestock and poultry genetic resources in China,is one of the indispensable livestock in Taihu Plain.It has a variety of excellent breeds characteristics,not only a large number of lambs,good wool production performance,the most important thing is the rapid growth and development of Hu sheep,meat production performance is ideal,is a rare variety of lambskin meat.With the change of sheep breeding mode,it is possible to artificially regulate the growth and development of sheep.Some feed additions can promote the growth and development of sheep.However,in recent years,the abuse of antibiotics in feed is prohibited in our country.Therefore,new preparations such as microecological agents,plant extracts and minerals that can promote the growth and development of livestock are widely used.Probiotics and plant essential oil feed additives have been widely concerned by researchers in improving the production performance of livestock.For this reason,in this study,biochemical indexes were determined by feeding compound probiotics and plant essential oil feed additives,and then the differentially expressed gene FGF18 was screened by transcriptome sequencing,and the effect of differentially expressed gene FGF18 on muscle growth of Hu sheep was further verified by molecular biology technology.This study provides a theoretical basis for exploring the molecular mechanism of adding compound probiotics and plant essential oils to improve the growth,development and meat quality of Hu sheep and gene regulation of sheep muscle growth.The results are as follows:1.Thirty-six male Hu sheep were randomly divided into 3 groups(compound probiotics group,compound probiotics + plant essential oil group and control group).There were 12 sheep in each group with a body weight of 20.00 ± 2.00 kg.The experiment period was 56 days,daily feed intake was recorded,feed weight ratio was calculated,jugular blood was collected,serum was separated,and biochemical indexes were detected.The results showed that the average daily gain of sheep in the probiotics + plant essential oil group was 135.86 ± 10.16 g,which was significantly higher than that in the control group(P < 0.01),and the feed weight ratio was the lowest among the three groups,about7.94 ± 0.42.On the 56 th day,the contents of total protein(TP),albumin(ALB),glutamic oxaloacetic transaminase(AST)and total cholesterol(TC)in the adding group were significantly higher than those in the control group(P < 0.05),and the content of ALB was significantly higher than that in the other two groups during the feeding period(P <0.01).2.Hu sheep fed with compound probiotics and plant essential oils were divided into two groups according to carcass weight,high meat yield group(n = 3)and low meat yield group(n = 3),a total of six.After slaughtering,the longissimus dorsi was collected and RNA was extracted and sequenced.Subsequently,the differentially expressed genes affecting meat production traits were analyzed by RNA-seq sequencing,and the differentially expressed genes most likely to affect muscle growth and development were selected for subsequent molecular level verification.The results showed that 80,714,496Clean＿Reads were obtained from the longissimus dorsi muscle of 6 sheep.According to GO enrichment analysis,2094 differential genes were obtained,including 1139 upregulated genes and 955 down-regulated genes.FGF18 with the most significant difference among the up-regulated genes was selected to quantitatively detect the tissue expression profile in 6 kinds of tissues.The results showed that the expression level of FGF18 in Hu sheep’s longissimus dorsi was significantly higher than that in other tissues(P < 0.05).The expression of FGF18 in high meat yield group and low meat yield group was detected by RT-q PCR.It was found that the expression of FGF18 in the longissimus dorsi of Hu sheep with high meat-yielding was significantly higher than that in lowyielding group.The protein expression was consistent with the results of m RNA(P <0.05).3.In order to further explore the relationship between FGF18 and muscle growth,FGF18 overexpression(pc DNA3.1-FGF18,pc DNA3.1-NC)and interference(sh RNAFGF18,sh RNA-NC)plasmids were constructed and transfected into the sheep primary myocyte.The expression of CDK4,cyclin-D1 and cyclin-D2,the key factors of cell proliferation,were detected by RT-q PCR.The proliferation of myocytes was detected by Ed U and CCK-8 analysis.The results showed that the expression of CDK4,cyclin-D1 and cyclin-D2 after overexpression of FGF18 was significantly higher than that of the negative control group,while the expression of CDK4,cyclin-D1 and cyclin-D2 showed the opposite trend after FGF18 inhibition(P<0.05).The results of Ed U and CCK-8 also proved that overexpression of FGF18 could effectively increase the proliferation of myocyte,while FGF18 inhibition would reduce the proliferation of myocyte.Summary,adding compound probiotics and plant essential oil to the diet can effectively increase feed intake,reduce feed-to-weight ratio and improve growth performance.Transcriptome sequencing results showed that FGF18 was an important candidate gene to regulate sheep carcass weight after feeding combined additives,and overexpression of FGF18 could promote sheep muscle cell proliferation.This study provides new ideas for improving sheep meat production performance and promoting the development of mutton sheep industry.