Long non-coding RNAs(lncRNAs) is a group of RNAs, which are longer than 200 nucleotides without containing functional open reading frames and cannot encode proteins. The studies indicate that lncRNAs can participate in the epigenetic modification. The most genome in mammals and other eukaryotes is transcribed in a developmentally regulated manner to produce large numbers of long non-coding RNAs. lncRNA can regulate DNA methylation, histone modifications, chromatin remodeling and other RNA interference through many ways, and the expression and role of lncRNA can be regulated by DNA methylation and chromatin remodeling. This article will review the lncRNAs function and its important role in gene regulation and medicine.

The analysis of different Chinese local pig breeds insulin promoter sequence provided the basis for studying miniature pig β cell function and preparing diabetes model. In this study, porcine insulin promoters（PIP）from three pig breeds(Tongcheng pigs, Guangxi Bama Miniature pigs and Wuzhishan Miniature pigs) had been amplified by PCR. By the comparative analysis, the sequences of local pig breeds characteristics were got. To verify the validities and specificities of the gene expression regulatory of the sequence, pancreatic tumor cells and transgenic mice were used. There were no differences among the three local pig breeds in 1.5 kb insulin promoter sequences. Compared with the sequence from foreign pig breeds, there were only three nucleotide differences. Dual luciferase reporter gene detection system proved that the promoter could efficiently start downstream gene expression; In transgenic mice with CHOP and UCP2, two target genes expressions in pancreatic tissue were significantly higher than that in the liver, back muscles and kidneys, and pancreas tissue protein abundance significantly increased. The result of this study shows that the insulin promoter of Chinese local pig breeds can regulate target gene expression in the pancreatic tissue, which lays the foundation for producing transgenic pig model of regulating β cell function.

This study was aimed to clone the porcine cyclin-dependent kinase 2 (Cdk2) gene and investigate the function of CDK2 encoded by Cdk2. In this study, porcine Cdk2 cDNA was cloned by reverse transcriptase-polymerase chain reaction (RT-PCR). Bioinformatic approaches were used to analyze the nucleotide and amimo acid sequences as well as the bio-function of CDK2. Semi-quantitative RT-PCR was used to analyze the tissue expression profile of Cdk2 mRNA, confocal microscopy was used to observe the subcellular localization of CDK2. Overexpression and shRNA techniques were both employed to investigate the role of CDK2 in cell cycle and cell proliferation regulation. Results showed that Cdk2 gene (GenBank accession no. JX967576) contained an 897 bp open reading frame and shared 94.2%, 94.0%, 93.8%, 93.4%, 91.8%, 91.0%, 90.6% and 89.9% identity with those of sheep, cattle, goat, human, golden hamster, house mouse, Chinese hamster and norway rat, respectively. Porcine Cdk2 gene coded 298 aa, with a molecular weight of 34 ku. The mRNA transcripts of porcine Cdk2 were found in 10 different porcine tissues by semi-quantative RT-PCR. Porcine CDK2 was localized in both the nucleus and cytoplasm and degradated through proteasomal pathway. Porcine CDK2 overexpression induced a significant decrease in proportion of cell at S-phase and G2/M-phase arrest, but no changes at G0/G1-phase. In contrast, when the CDK2 expression was down-regulated by shRNA, a significant decrease in proportion of cell at S-phase and G0/G1-phase arrest was observed, without changes at G2/M-phase. The porcine Cdk2 gene was successfully cloned and its function was also investigated.

 The intramuscular preadipocytes culture model in vitro of goat was established in this study, in order to research marker gene expression in intramuscular adipose tissue development and fat deposition in goat. The longissimus muscle of 1-day-old lamb was collected in this experiment, and digested by type II collagenase for 1.5 h, then filtered by 80 mesh, 400 mesh in sequence, after centrifugation, proliferative intramuscular preadipocytes were obtained using differential adhesion method. The morphological changes were observed, and fat content in cells was detected by oil red O staining. The genes expression of LPL, PPAR, FTO and LIPIN was detected using real-time quantitative PCR. The results showed that: the primary cultured cells were homogeneous, and the shortening-shaped or prismatic cells were showed in adherent growth, with the morphological characteristics of the intramuscular preadipocytes. The mRNA of FTO gene had a higher expression level in early stage, and the peak occurred at the 5-7 d in differentiation, and thereafter maintained a high level, then expression amount significantly decreased till 11 d (P<0.05). LIPIN mRNA fluctuated after the addition of inducer. PPAR mRNA maintained a relatively stable level in whole differentiation and proliferation process. LPL mRNA was expressed highly in the early stages of differentiation, and the expression decreased gradually with the increase of the degree of differentiation. The primary culture method of intramuscular preadipocytes in goat was established successfully in this research, and proliferation and differentiation process were reproduced in vitro. The amount of gene expression by quantitative analysis was: FTO> LPL> PPAR> LIPIN. This research laided the basis for further study of the fat deposition mechanism of intramuscular adipose tissue and improving the quality of goat.

To detect the location of ERα, ERβ and PR and the expression of ERα, ERβ and PR mRNA in the uterus of Jining Grey goats during postnatal development. In the uterus of different group, the SABC methods were used to study the distribution of ERα, ERβ and PR, and the real time fluorescent quantitative PCR were used to research the relative expression of ERα, ERβ and PR mRNA with GAPDH as an internal gene. ERα，ERβ and PR were malinly localized in the nuclei of the luminal epithelium, glandular epithelium, stromal cells, endothelium and smooth muscle cells of the uterus，and occasionally in the cytoplasm. The trends of the optical density in the immunopositive cells of ERα, ERβ and PR were similar with that of the expression of ERα, ERβ and PR mRNA. But the level of the expression of ERβ was less than that of ERα. From the birth day to the 30th day, the expression of ERα and ERβ remained at a lower level then increased gradually on the 60th day, ERα and ERβ reached its peak from the 90th day to the 120th day, and afterwards maintained at a high level. While the expression of PR was higher on the birth day, than reached its lowest level on the 30th day and significantly increased after puberty, and two peaks were found during the sexual maturity and maturation. ERα, ERβ and PR have an important role in regulating the proliferation and differentiation of the endometrial and the myometrial.

 The objective of this study is to analyze the reproductive hormone seasonal variations of year-round and seasonal estrous sheep. E₂, P₄, FSH and LH concentration in serum were determined, and the results could provide theoretical basis of hormone for regulating the seasonal reproduction in sheep. Blood samples were collected from five adult Small-tailed Han ewes and five adult Tan ewes in spring, summer, autumn and winter, respectively. The serum E₂, P₄, FSH and LH were measured by radioimmunoassay. According to the observation, the Small-tailed Han sheep kept estrous status during four seasons, while the Tan sheep started estrus in August and ended in March. The results showed that these four hormone level in Small-tailed Han sheep had no significant changes around the year. However, it was different in Tan sheep, P₄, FSH and LH concentration in estrous season were significantly higher(P<0.05) than those in anestrous season. According to the analysis between two breeds, E₂ level has no significant difference（P>0.05）around the year. The serum P₄ and FSH concentration in Small-tailed Han sheep were significantly higher(P<0.05) than in Tan sheep during spring and summer. The LH level in Small-tailed Han sheep was higher than in Tan sheep, however, which has no significant difference (P>0.05) between two breeds except in winter. These results indicated that a certain level of E₂ and P₄ were needed for sheep year-round estrous, and high level of FSH and LH were necessary for year-round estrus or polyembryony in sheep.

The present study was aimed to establish a novel method that can efficiently isolate and purify spermatogonial stem cells (SSCs) from goat testis. Single cell suspension was prepared from testis of 4-month old goat by three-step enzymatic digestion. The SSCs were enriched from the suspended cells via differential attachment in gelatin, collagen, and laminin coated dishes respectively, according to the differential adherence capability in different kinds of cells. Immunofluorescent staining by using SSC molecular marker PLZF and CDH1 was performed for analysis of enrichment efficiency of the SSCs. The percentages of SSCs were calculated through PLZF- and CDH1- positive cells. The enriched cells were primarily cultured and carried out identification. 1.36×10⁷ suspended cells were harvest from per gram of goat testis. 8.7×10⁴ cells were obtained after purification, in which the purity of SSCs was 87.0%. The purified cells could be cultured to form clusters in vitro, and CDH1 and PLZF were identified to express in the cells by immunofluorescent staining. The results show that a technique for purification of goat SSCs has been established by three-step enzymatic digestion and differential attachment. Goat SSCs purified by this technique can be cultured in vitro for proliferation and forming clusters.

The polymorphisms of MBL2 gene and their correlation to semen quality and progeny performance were investigated in order to find markers for semen quality and progeny performance in Chinese Holstein bulls. The polymorphisms of MBL2 gene were analyzed by the method of PCR-SSCP and sequencing in 218 adult Chinese Holstein bulls. The associations between mutation sites of MBL2 gene and semen quality and progeny performance were analyzed. The results showed that, there were five mutations, g.101 G/A, g.135 C/A, g.136 G/A and g.147 T/C in exon 1 and a novel SNP g.32 C/T in exon 2 (GenBank accession number: KC 832932). The g.101 G/A and g.135 C/A resulted in 31 Arg→31 Gln and 42 Pro→42 Gln, respectively. The site of g.101 G/A was significantly associated with sperm density (P＜0.05); the g.135 C/A was significantly associated with sperm motility and sperm density (P＜0.01); the g.136 G/A and g.147 T/C SNPs had very significant effect on sperm motility, sperm density, estimated breeding value of milk ( EBVM), estimated breeding value of fat (EBVF), estimated breeding value of protein ( EBVP) and estimated breeding value of somatic (EBVS) (P＜0.05); The g.32 C/T SNP had very significant effect on EBVS (P＜0.01). Five haplotype combinations were identified in exon 1 and the individuals with MN haplotype combination share the highest sperm motility, sperm density, EBVM, EBVF and EBVP to be positive for semen quality and dairy herd milk performance. The results indicate that the polymorphisms of MBL2 gene is correlated with semen quality and progeny performance in Chinese Holstein bulls.

The relationship between GDF-9 gene expression and expansion of COCs during in vitro maturation was investigated in this study. The expression of GDF-9 in bovine COCs during in vitro maturation(IVM)( 0, 6, 12, 18, 24 and 28 h) were detected by fluorescent quantitative RT-PCR method. The cumulus expansion was also observed. The RT-PCR result showed that GDF-9 mRNA expressed regularly in bovine COCs during in vitro maturation, which increased from 0-12 h culture and attained the highest expression at 12 h maturation culture, but subsequently decreased gradually and attained the lowest expression at 24 h maturation culture. The cumulus cells of COCs began to expand apparently at 12 h IVM culture, and increased expansion with maturation culture afterwards.The rusults showed that cumulus cells begin to expansion when expression of GDF-9 mRNA was highest in COCs, which suggested that GDF-9 may play essential role in cumulus expansion and oocyte maturation of bovine.

The purpose of this study was to clarify differential pattern of mRNA expression for genes related to different developmental stages of yak oocytes and preimplatation embryos, and to provide molecular evidence for gene expression of preimplatation embryonic development. The mRNA differential display reverse transcription PCR was used to identify differential expression bands for sequencing and alignment. The levels of mRNA relative expression were evaluated by RT-PCR in oocyte, 2-cell embryo, 8-cell embryo and blastocyst in yak. The results showed that three differential expression bands were highly homologous with zinc finger MYND-type containing 11 (ZMYND11), ribosomal protein L27a (RPL27A) and nucleosome assembly protein 1-like 1 (NAP1L1), respectively, except that one gene during morula stage showed no homology to any other known genes in GenBank. The relative expression levels of ZMYND11 genes increased in matured oocyte, 2-cell embryo, 8-cell embryo and in the blastocyst, with no significant differences between these stages (P>0.05). The relative expression levels of NAP1L1 genes were relatively low in matured oocyte, 2-cell embryo and blastocyst, with sharp increase in 8-cell embryo (P<0.05). The relative expression levels of RPL27A genes decreased during development, from matured oocyte to 2-cell, then to 8-cell embryo, and increased significantly in the blastocyst (P<0.05). ZMYND11, RPL27A and NAP1L1, the mRNA expression levels of which showed differences in different development stages of yak oocytes and preimplantation embryos, might be related to normal development and different physiological activities of preimplantation embryo in yak.

To study the effect of nano-ceria on production performance, nutrient digestibility, digestive enzyme activities and digestive enzyme gene expression of laying hens, the experiment was designed to preliminarily discuss the mechanism of improving production performance and nutrient digestibility of the optimum addition of nano-ceria. 396 26-week-old healthy commercial laying hens were selected and allotted randomly to four treatments, each treatment was represented by eight replicates of twelve each, and were fed the corn-soybean basal diet with 0, 30, 60, 90 mg·kg^-1 nano-ceria, respectively. The test lasted for seven weeks. The result showed that:(1)The nano-ceria had no significant effect on laying rate and feed egg ratio (P>0.05), but had significant effect on average egg weight, feed intake and broken rate of eggs(P<0.05). 30 mg·kg^-1 group had higher egg weight than other groups, 60 mg·kg^-1 group had lower broken rate of eggs than other groups. The nano-ceria had significant effect on calcium utilization ratio(P<0.05), but had no significant effect on the utilization rate of fat, crude protein, energy and phosphorus (P>0.05). (2)The nano-ceria had no significant effect on pancreatic lipase activities in intestinal(P>0.05), but had significant effect on pancreatic lipase activity in pancreas, the highest in 60 mg·kg^-1 group. The nano-ceria had significant effect on trysin and pancreatic amylase activity in intestinal and pancreas, the highest in 30 mg·kg^-1 group. (3)The nano-ceria supplementation had effect on mRNA level of digestive enzyme gene. Compared with the control group, 30 mg·kg^-1 group up-regulated mRNA level of trypsinⅡ gene (P<0.01), pancreatic amylase gene and trypsinⅠgene (P<0.05), but the change of mRNA level of pancreatic lipase was not significant(P>0.05). Compared with the control group, 60 mg·kg^-1 group up-regulated mRNA level of pancreatic lipase,but down-regulated mRNA level of trypsinⅡgene (P<0.01) and pancreatic amylase gene (P<0.05). Compared with the control group, 90 mg·kg^-1 group down-regulated mRNA level of trypsinⅡgene (P<0.01) and pancreatic amylase gene (P<0.05), but had no influence on mRNA level of pancreatic lipase gene (P>0.05). The result indicated that nano-ceria could improve production performance by improving digestive enzyme activities and nutrients apparent digestibility caused by activation of digestive enzyme system. Compared with the control group, 30 mg·kg^-1 group up-regulated mRNA level of trypsinⅡ gene, pancreatic amylase gene and trypsinⅠgene (P<0.05)，and then improved the egg weight and calcium digestibility; Compared with the control group, 60 mg·kg^-1 group up-regulated mRNA level of pancreatic lipase, down-regulated mRNA level of trypsinⅡgene and pancreatic amylase gene (P<0.05), the pancreatic amylase, trypsin activity and broken egg rate decreased; Compared with the control group, 90 mg·kg^-1 group down-regulated mRNA level of trypsinⅡgene and pancreatic amylase gene (P<0.05), so egg weight, feed intake and trypsin activity decreased, broken egg rate increased significantly. Comprehensive assessment, the diet with addition of 30 mg·kg^-1 nano-ceria was better.

The aim of this study was to explore the quantity and distribution characteristics of functional microorganisms in gastrointestinal tract of Liuyang Black goats, and provide foundation for further study on nutritional metabolism of these microbes. Microbes with known definite functions, such as bacteria and methogen, four cellulolytic bacteria, probiotics (Lactobacillus and Bifidobacterium), Prevotella ruminicola were selected in this study. Four Liuyang Black goats (1.5-year-old, (20.0±1.2)kg weight) were slaughtered at the same time and the digesta in the duodenum, jejunum, ileum, cecum, colon, rectum and rumen were collected for the isolation of the DNA, followed by the construction of the recombined standard plasmids. qRT-PCR was used for the absolute quantification of the microbes. The standard plasmids and curves of the microbes and methogen were constructed successfully and could be applied to the research. There were significant differences (P<0.01) in microbes between the 7 sections of the gastrointestinal tract of Liuyang Black goats. And the numbers of the microbes in the same section differed from each other significantly. Apart from the Butyrivibrio fibrisolvens, the largest number of the microbes was found in the rumen. In the intestinal tract, the number of microbes in the duodenum and jejunum was the least, while the number of microbes in the cecum, colon and rectum was the largest. The quantity and distribution characteristics of functional microorganisms in different gastrointestinal tracts of Liuyang Black goats significantly differ from each other (P<0.01). Overall, the number in the rumen was the biggest; the number in the cecum, colon and rectum was middle, followed by the ileum, while the least number was found in the duodenum and jejunum. This difference in the quantity and distribution characteristics of functional microorganisms in gastrointestinal tract has paved a way for the further study on different nutritional metabolism of these microbes.

This experiment was conducted to evaluate the effects of sodium butyrate on weaning stress of calves. A total of 24 healthy 31-day-old China Holstein calves were randomly allocated into 2 groups (control group and experimental group) with 12 calves per group. Calves in control group were reared by the feeding method of the farm, and the others in experimental group were fed diet supplemented with 1% sodium butyrate（dry matter basis）. All the calves were weaned at 60 d. The experiment lasted 45 days. The results showed as follows: 1) Compared with the control group, the live weight of calves in experiment group was increased by 3.86% (P<0.05) and 4.58% (P<0.01) at the age of 60 and 75 d; On the age of 31-60 and 61-75 d, the average daily gain (ADG) of calves in experiment group was increased by 6.33% (P>0.05) and 10.61% (P<0.01). 2) Compared with the control group, the contents of triglycerides, urea nitrogen, CRP, HP, IL-6, TNF-β in serum of calves in experimental group were significantly lower (P<0.01) and glucose was significantly lower (P<0.05) at weaning day; the contents of IgM of calves were significantly higher in experiment group than that in control group (P<0.05); the serum level of triglycerides, urea nitrogen, CRP, HP and cortisol of calves were significantly lower in experiment group than that in control group at 15 d after weaned (P<0.01) and glucose was significantly lower(P<0.05). 3) The rumen villous length and width was significantly higher in experiment group both at weaning day and 15 d after weaned (P<0.01). Compared with the control group, the villous length, crypt depth and V/C value of duodenum and jejunum were improved at different degrees. In conclusion, sodium butyrate supplementation can improve the growth performance, reduce the serum acute phase proteins and cortisol levels, and improve the immune function of weaning calves. Furthermore, it promote the development of the rumen and intestinal. In short, sodium butyrate alleviate weaning stress of calves.

The trial was conducted to evaluate the effects of different dietary protein and fat sources on nutrient digestibility and nitrogen metabolism of female blue foxes during fur development period. The animals were randomly assigned to four treatment groups with 10 females in each group. The randomization was based on age and initial body weight. The method of double-factor design is adopted. Two dietary protein sources were plant protein and animal protein (P plant and P animal ), two dietary fat sources were bean oil and lard (F plant and F animal), in four kinds of experimental diets. The trials of metabolism and nitrogen balance were used to analyse the digestibility of protein，digestibility of fat，nitrogen retention and protein biological value. The results showed as follows: digestibility coefficients of protein, ether extract, energy(P<0.01) and dry mass(P<0.05) in P plant group were significantly higher than that in P animal group, but nitrogen retention, net protein utilization and biological value of protein in P plant group were extremely significantly lower than that in P animal group(P<0.01); digestibility coefficients of protein, fat and energy in F animal group (lard group) were lower than that in F plant group (bean oil group), but lard could materially improve biological value of protein and materially reduce urinary nitrogen (P<0.01). In conclusion, nutrient digestibility of dietary plant protein source is higher, but biological value of protein is significantly lower than that of animal protein source. If feeding blue fox with plant protein mainly in practical production, it is should be considered the balance of amino acids. Fat digestibility of bean oil is higher, but lard can improve nitrogen metabolism and nitrogen retention. So the effect of production will be better when using mixed fat.

In order to investigate the epidemiology of Bartonella henselae infection, the 17-kDa protein gene of B. henselae was cloned into a prokaryotic expression vector (pET-28a) and constructed a recombinant plasmid pET-28a-17kDa. Then the pET-28a-17kDa was transformed into competent E. coli strain (BL21). The 17-kDa protein was expressed with IPTG induction and purified with nickel-agarose by affinity chromatography. Moreover, an indirect ELISA assay was developed to detect anti-B. henselae antibody using the purified protein as coating antigen. The optimized reaction conditions were as follows: antigen protein was coated at 37 ℃ for 2 h with 0.4 μg·mL^－1 concentration and then stored overnight at 4 ℃; The plates were blocked with 1% BSA at 37 ℃ for 3 h; The serum sample was incubated at 37 ℃ for 1 h with 1:100 dilution and secondary antibody was incubated at 37 ℃ for 0.5 h with 1∶15 000 dilution. The cutoff values of positive and negative results were OD_{450 nm} ≥ 0.392 and OD_{450 nm} ≤ 0.332 6, respectively. The specificity results showed that the protein could not react with positive sera of other six diseases (PRRSV, PCV2, CSFV, PRV, FMDV, M. suis) and the repeatability of intra- and inter-assay were excellent. Furthermore, total of 379 clinical sera samples obtained from swine farms were detected and the positive rate was 51.72%. In conclusion, the results showed that the indirect ELISA could be used for epidemiological surveys and diagnosis on Bartonella henselae.

MIC3 has high immunoreactivity, and plays an important role in recognition, adhesion and invasion for Toxoplasma gondii to host cells. To prepare the monoclonal antibody (McAbs) against recombinant MIC3 (rMIC3) of T. gondii, we expressed rMIC3 in Escherichia coli, and purified rMIC3 was immunized to BALB/c mice. B lymphocytes hybridization technique was applied to prepare the aimed McAb. Positive clones were screened with ELISA, and then were subcloned to establish stable cell lines. Ascites were induced to produce the McAbs, and its specificity was identified by Western blot analysis. The gold immunochromatographic lateral flow assay system was developed. The results indicated that two stable hybridoma cell lines (B7 and D3) were obtained. The McAbs subclasses all belong to IgG2a and their ELISA titers of the ascite fluid were 1:640 000 and 1:1920 000, respectively. Western blotting analysis confirmed that the two McAbs all can identify the protein MIC3 from lysed tachyzoites and purified rMIC3. Purified polyclone antibody D3 was labeled with colloidal gold particle, and the minimum detectable limit of tachyzoite was 4×10⁴ CFU·mL^－1. This study laid the foundation for further study on related applications.

The present study was conducted to prokaryotically express the nucleocapsid (N) protein of Schmallenberg virus (SBV) and prepare its polyclonal antibodies. Using the total RNA of SBV (isolate BH80) as a template, RT-PCR was utilized to amplify the full-length coding sequence of the N gene, which was then cloned into the prokaryotic expression vector pET-28a-c(+) to construct the recombinant plasmid pET-28a-SBV-N. After verification using the double-enzyme cleavage method and sequence analysis, the recombinant plasmid pET-28a-SBV-N was transformed into the competent E. coli BL21 (DE3) cells which were then induced by isopropyl-β-thiogalactopyranoside (IPTG). The recombinant His-SBV-N fusion protein was purified under native conditions using a nickel ion metal chelate column. Subsequently, the purified His-SBV-N protein was used as an immunogen to immunize the New Zealand white rabbits to prepare its polyclonal antibodies. After purification of the prepared polyclonal antibodies using the immunoaffinity chromatography method, Western blot analysis and indirect immunofluorescence assay were used to validate the polyclonal antibodies. Our results demonstrated that: (1) The recombinant His-SBV-N fusion protein was efficiently expressed in E. coli BL21(DE3) in the form of both soluble expression and inclusion bodies, and the molecular weight of His-SBV-N protein was about 30 kDa; (2) The titer of the prepared polyclonal antibodies was 1: 64 000. The prepared polyclonal antibodies not only can specifically react with the recombinant His-SBV-N fusion protein, but it also can recognize different isolates of SBV. However, the prepared polyclonal antibodies also have cross-reactivity with the N protein of the most genetically related viruses, such as Shamonda virus, Douglas virus and Akabane virus, within the family Bunyaviridae. Undoubtedly, the successful preparation of both His-SBV-N fusion protein and its polyclonal antibodies lays a material foundation for the establishment of serological methods for SBV detection.

The objective of this study was to prepare monoclonal antibody against glycoprotein C (gC) protein of infectious bovine rhinotracheitis virus (IBRV) and to analyze its biological characteristic. BALB/c mice were immunized with purified recombinant gC protein expressed by E. coli and the spleen cells of immunized mouse were fused with SP2/0 myeloma cells. An indirect ELISA using the purified gC protein was developed to screen positive antibody-producing cells. A hybridoma stably secreting MAb designated as 3D6 was obtained against gC protein. The monoclonal antibody belongs to IgG1, and the light chain was κ. The titers of supernatant and ascites were 6.4×10³ and 1.28×10⁵ as detected by ELISA, respectively. The result of western blot assay and IFA suggested that the MAb could recognize IBRV. A sandwich ELISA was established by using 3D6 secreted monoclonal antibody. The results indicated that the ELISA possessed good specificity and higher sensitivity. The monoclonal antibody could be used to diagnose IBRV and further research gC protein function.

Porcine eperythrozoonosis (PE) caused by Mycoplasma suis (M. suis) infection is a febrile acute icteroanemia in pigs and human. In order to study the pathomechanism of PE, clinical symptoms, hematological parameters, M. suis loads, cellular and humoral immunity, gross pathology and histopathology were system recorded to monitor the process of this disease in models of splenectomized and non-splenectomized pigs. The result showed that the M. suis loads in splenectomized pigs maintain higher level (1.0×10⁸ copies per mL) and longer time than that of non-splenectomized pigs. Correlations between M. suis loads and the values of WBC, hematocrit, RBC and Fe were significant. The antibody against MSG1 was detected in only one pig, whereas the IL-4 and IFN-γ were not detected in all of the groups during the whole experiment. PE was reproduced and the process of this disease was recorded in this study which plays a fundamental role in revealing the pathomechanism, providing the guideline of prevention and diagnosis of this disease.

The objective of the study was to explore whether progesterone meet the requirement for acting on neurons of Nodose Ganglion (NG), thereby involving in the regulation of gastrointestinal activity by affecting primary afferent neurons of viscera. We used immunohistochemical SP method to detect the expression of progesterone receptor (PR) in Nodose Ganglion of female goat. The results indicated that PR immunoreactive products distributed widely in Nodose Ganglion of female goat, varying degrees of which were shown in neurons, satellite cells, Schwann cells and nerve fibers. The cell membrane of neurons presented brown as strong positive, and the Nissl bodies was moderate or weak positive in the cytoplasm. Most of neurons’ karyoplasm was strong positive,but the nucleolus was clear vacuoles. PR was strong positive on the majority of satellite cells, but others’ nucleus revealed a vacuolization having no staining. There were flaxen PR-immunoreactive products performing weak positive in Schwann cells and nerve fibers. Image analysis shows that compared with other non-nerve cells, PR of neurons was extremely significant (P＜0.01). The result proved that the visceral sensory neurons in NG were the main target cells of progesterone. That is to say, progesterone possibly impacts on visceral sensory neurons in NG of female goats, thereby affecting the activities of gastrointestinal activity through afferent part of the visceral reflex arc, and the PR in NG acts as the network node to coordinate the endocrine regulation of progesterone and neuroregulation of autonomic nerve on gastrointestinal function.

To ascertain the effect of different culture conditions on diversity of endophytic fungi from Oxytropis kansuensis in China, the study collected Oxytropis kansuensis samples in Qilian county of Qinghai province, using the method of surface disinfection and different medium, to isolate and purify the endophytic fungi in stems and leafs, then the morphology techniques and ITS sequence analysis were used to identify their species, and calculated diversity index by Biodap. The results showed that 433 fungal colonies were isolated from 629 samples, which were combined to 28 kinds of different endophytic fungus. Undifilum oxytropis was the dominant strain, and advantage was more obvious in the light condition. The Shannon-Weiner index was 1.99, evenness index was 0.60, simpson’ diversity index (D) was 0.257. The diversity was different under different culture conditions. The Shannon-Weiner index was highest in the dark condition, and the simpson’ diversity index was biggest in the light condition. The results showed that the diversity and isolation rate of endophytic fungi from Qilian county of Qinghai province was different under different culture conditions. Dark condition and PDA medium could improve the isolation rate of endophytic fungi, dark condition and BDA medium can increase the diversity, as the Undifilum oxytropis, the light condition and BDA medium can increase the separation rate.

Through the application of biological NMR technique combines principal component analysis (PCA) and orthogonal partial least-squares discriminant analysis (OPLS-DA), changes of endogenous metabolites with clinical and subclinical ketosis cows were analyzed, which may help to study the effects of ketosis on cow metabolic process, and find the differences metabolites. The plasma metabolic profiles of the three groups were obtained by ¹H nuclear magnetic resonance (¹H NMR). Differences in metabolites were acquired by analyzing the collected data with mode analysis. Results were as follows: compared with the healthy control group, 23 different metabolites were obtained in subclinical (13 were up-regulated, 10 were down-regulated) and clinical (6 were up-regulated, 17 were down-regulated) ketosis groups. By comparison, 28 different metabolites were obtained between clinical ketosis group (4 were up-regulated, 24 were down-regulated) and subclinical ketosis group. The results indicate that ¹H NMR spectra combined with pattern recognition technique can effectively get the different metabolites of diagnosing clinical and subclinical ketosis, and panoramic reveal that widely metabolic disorder on cows in the process of ketosis, which lays the foundation for the exploration of ketosis mechanism and obtaining new biomarkers in the future.

A high-throughput and efficient Affymetrix rat genome array was used to investigate the pharmacological mechanism of the traditional Chinese medicine, Pulsatillae Decoction (PD), used for the treatment of diseases induced by lipopolysaccharide (LPS). Endothelial cell was challenged with 1 μg·mL^－1 LPS for 3 h, then treated with PD at a concentration of 1 mg·mL^-1 for 12 h. Total RNA of each treated group was extracted from cultured endothelial cell for detection by the Affymetrix Rat Genome 230 2.0 Array. The samples from LPS group and blank control group were subjected to Real-time RT-PCR, using the chimeric fluorescence method. The results showed that in LPS group, 36 genes were upregulated and 33 genes were downregulated in comparison with Blank group; In LPS-PD group, 566 genes were upregulated and 12 genes were downregulated in comparison with LPS group, and 93 genes were upregulated and 29 genes were downregulated in comparison with Blank group. The RT-PCR ratios for the target genes Il1α, Il6, Edn1, Tnfsf11 and Ptges were 9.78, 1.76, 1.43, 2.98 and 2.12 respectively, which were very similar to the ratios determined by the gene chips (10.57, 1.53, 1.59, 3.16 and 1.71, respectively). These results indicate that PD could simultaneously reduce the translated level of inflammatory factors, suppress the transcription of apoptosis genes, accelerate the protein synthesis of organism and enhance the energy metabolism of cells, thus play the united antiendotoxin action.

In order to explore the effects of polysaccharide extracted from 11 kinds of traditional Chinese medicines on immunomodulatory in different MHC B-LβⅡ genotype chickens, PCR-RFLP technique was applied to group one-day-old male liangfeng Green-Feet Ma chickens according to different MHC B-LβⅡ genotypes, then they were given high, middle and low dose of polysaccharides extracted from traditional Chinese medicines and saline, respectively. The effects of traditional Chinese medicinal polysaccharide extraction on antibody response kinetics to ND vaccine and IFN-γ and IL-4 contents in serum were determined by ELISA method. The results showed that: (1) Traditional Chinese medicinal polysaccharide extraction at different dose could improve ND antibody level, IFN-γ and IL-4 contents in each MHC B-LβⅡ genotype chicken, and in high dose traditional Chinese medicinal polysaccharide extraction, AA genotype chicken were higher than that of AB, BB genotype, in middle and low dose traditional Chinese medicinal polysaccharide extraction, AB genotype were higher than that of AA, BB genotype; (2) The ND antibody level and IL-4 contents of AA genotype chicken in high dose traditional Chinese medicinal polysaccharide extraction were higher than that in middle and low dose group, the ND antibody level, IFN-γ and IL-4 contents of AB, BB genotype chicken in middle dose traditional Chinese medicinal polysaccharide extraction were higher than that in high and low dose group. These results indicated that the polysaccharide extracted from 11 kinds of traditional Chinese medicines could stimulate ND antibody production and cytokines IFN-γ, IL-4 excretion in different MHC B-LβⅡ genotype chickens, and the optimum immunomodulatory dose were different in each MHC B-LβⅡ genotype chicken.