With the application and development of the next generation sequencing technique，the researches of genomics are constantly updating，which finds out new solutions and technologies to genomics.The genome sequencing is competent to learn the population evolution，gene composition and gene regulation deeply，especially the application and development of genome resequencing technology，which makes the genome research come into being a new era in multiregion，diversification and multifunction.Nowadays the next generation sequencing technique has made a large progress in mutation detection，fine mapping of important genes，genetic map construction，analysis of population evolution，and so on.The review states application status and development tendency of whole genome sequencing technology and reduced-representation genome sequencing technology in animal genome resequencing.

With the development of animal molecular reproduction and embryo engineering technology，the production of genetically superior offspring has become a reality.The embryo engineering technology of animal embryo reproduction has been becoming an important technical approach in animal reproduction fields.The studies of animal embryo breeding include：1) the selection and optimization of embryo engineering technology in animal breeding program；2) the establishment of different systems of animal embryo breeding and the solution exploration of the issues of animal breeding such as inbreeding avoidance；3) the scientific assessment of genetic progress and biosafety in animal embryo breeding.The applications of these technological strategies in animal embryo breeding can be summarized as following：the purebred breeding through expanding group of outstanding individuals；the breeding of new varieties by transgene or gene modification technique；the breeding conservation by using animal germplasm resources.The technology of animal embryo breeding is the development and improvement of the traditional animal breeding theories，as well as the reflection of animal embryo biotechnology application in real animal production.This animal embryo breeding technology has broad application prospects in accelerating the development of animal breeding，the breeding selection，and cultivating new varieties.

Pasteurella multocida (P.multocida)，a Gram-negative cocco-bacillus，facultative anaerobic bacterium，is the causative agent of serious diseases in a wide range of animals.Lipopolysaccharide (LPS) is an important virulence factor，and is also a major immune protective antigen of P.multocida which is currently classified into 16 Heddleston serovars based on LPS.Unlike the other Gram-negative pathogens，P.multocida LPS is only made up of lipid A and core oligosaccharide，lacking an O-antigen.Recently，chemical structures and biosynthesis genes of 16 Heddleston serovars LPS have been determined by using mass spectrometry and gene sequencing.The inner core structures are highly conserved among different serovars，and genes required for the assembly of the inner core are located in several regions of the genome.In contrast，the outer core structures are distinct，and genes required for the biosythesis of the outer core structures are clustered in a single locus between the conserved genes priA and fpg.Some LPS serovars genetically have relationship each other，sharing the same outer core biosythesis locus，but producing different LPS molecules due to mutations within glycosyltransferase genes.Furthermore，LPS structures of P.multocida are related to bacterial virulence.Here，we summarize LPS structures，biosythesis genes and the relationships between LPS and bacterial virulence in P.multocida.

In order to understand the transcriptional regulatory mechanism of ssc-miR-148a，we have cloned and analyzed its promoter.Firstly，we designed specific PCR primers to amplify 3 upstream fragments of ssc-miR-148a precursors，then inserted them into pGL3-Basic expression vector.Based on the amplified fragments，ssc-miR-148a promoter region，methylation sites and transcription factor binding sites were predicted by bioinformatics.The plasmid was transfected into 293T cells to analyze the promoter activity.Expression of ssc-miR-148a and DNA methylation transferase 1 (DNMT1) in treated porcine fibroblasts by Basic Fibroblast Growth Factor（bFGF）with different concentrations was detected.The results showed that the cloned 2 043 bp sequence had promoter activity，which had 5 CpGs and transcriptional factor binding sites，such as Sp1，AP2.After treated porcine fibroblasts with 0，5 and 10 ng•mL^-1 bFGF，the expression of ssc-miR-148a was decreased significantly（P<0.05），DNMT1 mRNA level increased significantly (P<0.05).The promoter activity significantly reduced (P<0.05)，but no significant difference between 5 and 10 ng•mL^-1 concentrations (P>0.05).The results indicate that the promoter region of ssc-miR-148a locate at －2 043 bp，which has Sp1 transcription factor binding sites.The expression of ssc-miR-148a is regulated by the bFGF.

In this study，we analyzed genetic differentiation between small- and large-sized Chinese indigenous pigs，aiming to detect signatures of selection and to identify candidate genes and mutations related to small body size.Fixation index (Fst) and heterozygosity were calculated based on whole-genome resequencing data from miniature pigs (Wuzhishan and Bamaxiang) and large-sized pigs (Jinhua，Erhualian and Hetao).The regions with larger Fst values and lower heterozygosity were regarded as candidate regions of selection.Through further annotation on gene function，pathway and mutations of the genes located in the candidate regions，we sought to screen genes and mutations related to body size.In total，32 475 218 SNPs were obtained after stringent filtering from whole-genome resequencing data.Fst and heterozygosity were calculated using 40 kb sliding window strategy.And 242 putative selection regions were kept by applying a Z-transformed Fst (Z(Fst))>5.There are 13 regions overlapped between these 242 regions and the 200 regions with lowest Z-transformed heterozygosity (Z(H))，encompassing 28 genes.Seventeen out of the 28 genes are found to be associated with growth or body size through PANTHER annotation.Among these genes，IGF1R and GUCY1A3 were the most promising candidate genes.Together，by applying population genetic methods，we can detect regions of signature of selection related to body size in pigs.And after comprehensive analysis in gene function and mutations，we proposed that mutation(s) associated with small-sized phenotype in Wuzhishan and Bamaxiang pigs might be located in the non-coding regions detected.

The objectives of this study were to investigate the developmental changes of Myogenin (MyoG) and Phosphotyrosine interaction domain containing 1 (PID1) mRNA expression in different parts of muscles in sheep and research its effect on intramuscular fat (IMF) accumulation.Five Aohan fine wool sheep were slaughtered at 2，4，5，6，12 months，respectively，to collect samples from longissimus dorsi and biceps femoris，for determining the IMF content.The mRNA expression of the MyoG and PID1 in 2 parts of muscles was investigated by real-time PCR，and association between their mRNA levels and IMF contents were also analyzed.The results showed that，2-5 month，with growing of the age，the IMF contents of longissimus dorsi and biceps femoris increased.And 5-12 month，the IMF contents remained unchanged.The IMF content of longissimus dorsi were extremely significantly higher than biceps femoris (P<0.01) at the same age.MyoG and PID1 mRNA expression level was different in 2 parts of muscles and had tissues-dependent.In longissimus dorsi， MyoG expression level was the highest (P<0.01) in 5 month old sheep，PID1 expression level was the highest (P<0.05) in 6 month old sheep，there was an increase and then decrease trend in the 2 gene expression.In biceps femoris，the MyoG and PID1 expression level difference was extremely significant(P<0.01) among different month ages，and MyoG expression level presented increased and declined slowly with growing，PID1 expression level presented declined-increased with growing.In the same month，MyoG and PID1 expression level had significant difference in different parts.Correlation analysis showed that MyoG and PID1 expression level was inordinately positively related to IMF content.In conclusion，MyoG expression has positively effect on IMF accumulation in sheep，and PID1 expression is likely to influence on IMF accumulation.

The aim of this study was to acquire key molecular marker related to inflammatory reaction of bovine mammary epithelium cells induced by Methicillin-resistant Staphylococcus aureus (MRSA).In this study，bovine mammary epithelium cell line (Mac-T) was infected with MRSA in vitro by compared with the cells treated by S.aureus as well the untreated cells.The expression levels of two candidate genes associated with subclinical mastitis ((trafficking protein particle complex 9(TRAPPC9)，Janus Kinase 2(JAK2)) and one pro-inflammatory gene (interleukin 6，IL-6) were detected and compared among infected groups and non-infected group at different time points.Comparing with the untreated cells，the results showed that the mRNA expression level of TRAPPC9 significantly decreased (P<0.05) in the Mac-T cells at 6 h post stimulation by MRSA，while that of JAK2 significantly decreased (P<0.05) at 8 h；And the expression levels of TRAPPC9 and JAK2 in the cells significantly decreased (P<0.05) at 6 h post infection by S.aureus.Compared with S.aureus stimuli，the expression levels of TRAPPC9，JAK2 and IL-6 in the Mac-T cells were moderately higher at 6 and 10 h post infection by MRSA.Especially，the expression level of TRAPPC9 was significantly higher in MRSA treated cells (P<0.05) compared to S.aureus treated cells at 6 h post infection.These results suggest that the genes expression changes in the Mac-T cells were similar after the infection of MRSA and S.aureus，while the gene expression changes of JAK2 in the cells was delayed from MRSA infection compared to S.aureus.TRAPPC9 can be used as the preferred molecular marker for detecting MRSA and S.aureus infection in bovine mammary epithelium cells.

To reveal the molecular mechanism underlying in vitro maturation of the buffalo oocyte and select the proteins associated with the oocyte maturation quality，sodium dodecyl sulfate polyacrylamide gel electrophoresis coupled to reverse-phase liquid chromatography tandem mass spectrometry (RP-LC-MS/MS) proteomics technology were applied to identify the buffalo oocyte proteome at germinal vesicle (GV) stage and metaphase II (MII) stages.In total，647 and 570 proteins (FDR<1%) were identified from GV and MII stage buffalo oocytes，respectively.Among them，414 proteins were common identified，233 and 156 proteins were uniquely identified，respectively.The semi-quantitative information of the identified proteins was obtained using spectral counting method.Nine high abundance proteins were identified from GV and MII stage buffalo oocytes，eight of which were common proteins，including MVP，GATM3，PADI6，PRDX2，NE2，DYNLL1，AKR1B1 and LOC786101.HSP90α and GSTP1 were unique high abundance proteins identified in GV and MII stage buffalo oocytes，respectively.It indicated that they might be closely related to buffalo oocyte maturation.Results of GO (Gene Ontology) and KEGG analysis showed that these common identified proteins involved in metabolism pathways，such as pyruvate metabolism，glycolysis，tricarboxylic acid cycle (TCA)，and pentose phosphate pathway，etc，which suggested that high metabolism level was needed to complete meiotic maturation in buffalo oocytes.Proteins involved in oxidative phosphorylation，ribosome，and proteasome pathway were uniquely identified in GV stage oocytes，indicating that high level of oxidative phosphorylation and protein synthesis might exist in GV stage oocyte，and the proteasome pathway might be essential for the maturation progression of buffalo oocytes.Proteins involved in DNA replication，amino sugar and nucleotide sugar metabolism pathway were uniquely identified in MII stage oocytes，demonstrating that the oocytes might need to prepare genetic material for the fertilization and early cleavage divisions.Overall，above results revealed the protein expression profile of buffalo oocytes before and after maturation，they are useful to understand the molecular mechanisms underlying buffalo oocyte maturation.

This study aimed to investigate expression of toll-like receptors in different segments of intestinal tract in duck at the peak and end of lay.Twenty laying ducks at the peak of lay were randomly assigned to 2 groups.One group ducks were given a restricted feed for 7 days，and then provided with feed ad libitum for 10 days.Another group ducks were provided with feed ad libitum for 17 days.Expression of TLR 1-1，TLR1-2，TLR2-1，TLR2-2，TLR3，TLR4，TLR5，TLR7，TLR15 and TLR21 were analyzed in duck duodenum，jejunum，ileum and caecum by RT-PCR and qRT-PCR methods.The results showed that 10 TLRs genes were all expressed in duodenum，jejunum，ileum and caecum.Except for TLR1-2，TLR2-2，TLR4 and TLR21，the other TLRs genes mRNA expression was significantly different among different segments of duck intestinal tract (P<0.05).Of these，TLR3，TLR5 and TLR7 mRNA expression were higher in duodenum than in the other tissues，while TLR1-1，TLR2-1 and TLR15 mRNA expression were higher in caecum than in the other tissues.Except for TLR4 and TLR7，the other TLRs genes mRNA expression was higher in ducks at the peak of lay than that in ducks at the end of lay (P<0.05).In conclusion，a TLR-mediated innate immune response mechanism exists in the laying duck intestinal tract，and there was stronger ability to recognize pathogens in duck intestinal tract at the peak of lay than that at the end of lay.

 In order to detect the relationship between polymorphism of MC1R gene and coat color in fox，a total of 163 skin samples of 12 coat color foxes were collected.The nucleotide sequence (1 054 bp) of fox MC1R gene were obtained by the method of PCR and direct sequencing，and the polymorphism were analyzed.The population genetics were analyzed using PopGen32 and SHEsis softwares.The effect of mutations on the function of MC1R gene was evaluated using PANTHER software.The relationships between the variable sites and coat color were analyzed by the statistical methods of SPSS bivarate correlation analysis.Two adjacent missense mutations (c.40A>C and c.41C>T) were found in the coding region of fox MC1R gene，which resulted in codon change of p.Thr14Ile or p.Pro14Leu.When 40 site was A，it led to the substitution between threonine (Thr) and isoleucine (Ile).When 40 site was C，it led to the substitution between proline (Pro) and leucine (Leu).All the genotypes of two coat color foxes belonging to Alopex were TT.However，the genotypes of most coat color foxes belonging to Vulpes were CC.It was supposed that 41 site was important in distinguishing Alopex and Vulpes.The in silico functional analysis showed that the amino acid substitution at p.Pro14Leu had significant impact on the function of MC1R.The statistical analysis showed the polymorphism of 41 site had significant low correlation with the fox coat color.The results indicate that SNP c.41C>T in the coding region of the MC1R gene is probably associated with the coat color in fox. In order to detect the relationship between polymorphism of MC1R gene and coat color in fox，a total of 163 skin samples of 12 coat color foxes were collected.The nucleotide sequence (1 054 bp) of fox MC1R gene were obtained by the method of PCR and direct sequencing，and the polymorphism were analyzed.The population genetics were analyzed using PopGen32 and SHEsis softwares.The effect of mutations on the function of MC1R gene was evaluated using PANTHER software.The relationships between the variable sites and coat color were analyzed by the statistical methods of SPSS bivarate correlation analysis.Two adjacent missense mutations (c.40A>C and c.41C>T) were found in the coding region of fox MC1R gene，which resulted in codon change of p.Thr14Ile or p.Pro14Leu.When 40 site was A，it led to the substitution between threonine (Thr) and isoleucine (Ile).When 40 site was C，it led to the substitution between proline (Pro) and leucine (Leu).All the genotypes of two coat color foxes belonging to Alopex were TT.However，the genotypes of most coat color foxes belonging to Vulpes were CC.It was supposed that 41 site was important in distinguishing Alopex and Vulpes.The in silico functional analysis showed that the amino acid substitution at p.Pro14Leu had significant impact on the function of MC1R.The statistical analysis showed the polymorphism of 41 site had significant low correlation with the fox coat color.The results indicate that SNP c.41C>T in the coding region of the MC1R  gene is probably associated with the coat color in fox.

 This experiment was conducted to study the effect of different patterns of farrowing pens on reproductive performance and stress level of sows，in order to explore a kind of welfare and friendly feeding mode of lactating sows that could be popularized and applied in practical production.Twenty-four hybrid sows (Large White × Landrace) which had same parity，similar pregnancy and body condition were randomly assigned to 3 treatments：farrowing crate＋high bed group (FCB，n=8)，freedom farrowing pen＋high bed group (FFPB，n=8) and freedom farrowing pen＋partially fermented bed surface group (FFPF，n=8).The 7^th day before farrowing，sows were transferred to different patterns farrowing pens.The 21^st day after farrowing，piglets were weaned.The results showed that：1) farrowing time in FFPB and FFPF was significantly lower than in FCB (P＜0.05)；average farrowing interval in FFPB was significantly lower than in FCB (P＜0.05)；the levels of blood plasma oxytocin (OT) and prolactin (PRL) in FFPB showed an increasing trend (P＜0.10).2) The 2^nd day after transferation of sows to farrowing pens，waist surface temperature in FFPB and FFPF was significantly lower than in FCB (P＜0.05).The 7^th day after farrowing，salivary α-amylase (AMY) level in FFPB and FFPF was significantly lower than in FCB (P＜0.05).The 14^th day after farrowing，salivary AMY level in FFPF was significantly lower than in FCB (P＜0.05)，and cortisol (COR) level in FCB showed increasing trend (P＜0.10).These results indicated that：at the day of farrowing，reproductive hormones levels in freedom farrowing pens were increased while farrowing duration and average farrowing interval were reduced.Within 1 to 2 weeks after farrowing，stress level in freedom farrowing pens was obvious decreased，indicating freedom farrowing pens were more close to the welfare and friendly feeding mode of feeding sows.

This experiment was conducted to investigate the effects of 4 LED spectral combination (named: A (21% Blue+30% Green+24% Yellow+25% Red)，B (35% Blue+35% Green+18% Yellow+12% Red)，C (27% Blue+30% Green+22% Yellow+21% Red) and D (42% Blue+28% Green+18% Yellow+12% Red)) on growth，immune function，carcass performance and welfare of AA broilers，aiming to provide theoretical and practical guidance for poultry production.A total of 600 1-day-old AA broilers were used.Each lighting treatment included 5 replicate groups with 30 chicks(15 male，15 female) per group.Feed and water were provided for ad libitum.Light program was 20 lx for the first week，and 5 lx (14L∶4D∶2L∶4D) till the end (6 weeks of age).The results showed that broilers reared under C light source significantly decreased average daily gain than A，B and D groups at 4-6 weeks by 14.45%，13.76% and 12.91%，respectively (P<0.05).C light source had a tendency to decrease average daily gain of 1-6 weeks (P=0.07).C light source significantly decreased the average daily feed intake of 4-6 weeks and 1-6 weeks compared to D light source (P<0.05).A light source significantly decreased feed gain ratio compared to D group by 9.14% (P<0.05).Antibody titers versus Newcastle disease (NDV) and avian influenza (Re-6) in D light source was significantly lower than that in other light groups (P<0.05).A significantly decrease of the gait score was observed in birds reared under A，C and D light source groups (by 44.97%，26.32% and 33.33%，respectively) compared to B light source (P<0.05).Spectral combination had no significant effect on eyeball development or carcass traits (P>0.05).It was concluded that A light source not only good for growth performance of growing period，but also increase the immune function and welfare of AA broilers.

The aim of this study was to examine the effects of oviposition and feeding behavior on core and skin temperature of laying hens and diurnal variation of core and skin temperature of laying hens.Twelve Jing Hong laying hens aged 34 weeks were assigned to 6 cages，one cage included 2 birds and were kept at environmental control chambers（ambient temperature：20 ℃，relative humidity：60%）.All hens were exposed to 16L：8D until the end of experiment and fed twice a day.Miniature temperature data loggers were used to measure the core and skin temperature，and the data was recorded every 3 minutes，which lasted 48 h.The time of oviposition and feeding were recorded by digital infrared video camera.The analysis results by repeated measurement showed as follows：1) Oviposition had effect on the core temperature(P<0.001) and the skin temperature (P=0.082).Thereinto，the core temperature of egg-laying period was 0.21 ℃ higher than that of pre-laying (P<0.01) and 0.37 ℃ higher than that of post-laying (P<0.01).Compared with the pre-laying period，the skin temperature of egg-laying period had a tendency to increase(P=0.058).2) Feeding had significant effect on the core(P=0.01) and skin temperature(P=0.006).Compared with the core and skin temperature of the 1 h before feeding，the core and skin temperature of the 1-2 h after feeding increased 0.12 ℃ (P<0.05) and 0.35 ℃（P<0.05），respectively；2 h after feeding，the core and skin temperature were back to normal.3) During the day（11：00-12：00，15：00-16：00，19：00-20：00），the core and skin temperature had no significant differences（P>0.05），which significantly higher than that during the night（23：00-24：00，03：00-04：00，P<0.01）.In conclusion，the oviposition and feeding behavior significantly improved the core and skin temperature.Therefore，evaluating the comfortable level on the basis of body temperature in laying hens，the interference of oviposition and feeding should be avoided.

The present study was conducted to investigate the effect of high-fat (HF) diet on meat quality traits and skeletal muscle proteome of growing-finishing pigs.Twenty growing-finishing pigs with an average initial BW of (73.28±0.57) kg were randomly allocated to control group (control diet，supplemented with 10.0% cornstarch) and treatment group (high-fat diet，supplemented with 10.0% lard) to study the effect of dietary fat content on growth performance，meat quality traits and skeletal muscle proteome of growing-finishing pigs.There were 10 replicates in each treatment group with 1 pig per replicate.The experiment lasted 42 d.The results showed that pigs fed the HF diet had a greater average daily gain (P<0.05)，but a lower ratio of feed to gain than pigs fed the control diet (P<0.01).The pH_{45 min}，pH_{24 h}，intramuscular fat content，lightness，redness，yellowness，cooking loss，drip loss and shear force in skeletal muscle of pigs were not affected by dietary fat content (P>0.05).Proteomic analysis revealed that pigs fed the HF diet had greater expression levels of calreticulin，myosin heavy chain，capping protein，pyruvate kinase，enolase，L-lactate dehydrogenase，triosephosphate isomerase，adenylate kinase，apolipoprotein A and B，fatty acid binding protein and heat shock protein 27 and 70 ku，but lower expression levels of myosin light chain and phosphoglucomutase than pigs fed the control diet.In conclusion，the HF feeding could improve the growth performance，and upregulate the expression levels of proteins related to lipid metabolism，glucose and energy metabolism in skeletal muscle of growing-finishing pigs.

The aim of this experiment was to study the effect of dietary nutrient levels on the number of related microbes，including Succinivibrio dextrinisolvens(S.dextrinisolvens)， Prevotella ruminicola(P.ruminicola)，Selenomonas ruminantium(S.ruminantium)，Butyrivibrio fibrisolvens(B.fibrisolvens)，Ruminococcus flavefaciens(R.flavefaciens)，Veillonella parvula(V.parvula） and Methanogens，pH and VFA levels in the rumen of Tan sheep aged from 150 to 180 days.One hundred and twelve 105-day-old Tan sheep（half males and half females） with the similar average body weight (BW) of (29±1.25) kg and better body condition were allotted randomly to 4 dietary treatments (group Ⅰ，Ⅱ，Ⅲ and Ⅳ).Each treatment had 4 replicated pens，with 7 sheep in each pen.From group Ⅰ to Ⅳ，energy of diets were 7.08，8.09，9.10 and 10.11 MJ• kg^-1，respectively；dietary protein levels were 8.04%，9.19%，10.34% and 11.49%，respectively.The ratio of energy to protein was kept unchanged (about 0.88).The experiment lasted for 75 days，including 15 days for adaptation.On 150 and 180 days of age, one sheep was randomly selected in each replicate for slaughter and rumen contents were collected，pH was tested and VFA levels in the rumen were analyzed by gas chromatography measurement.Genomic DNA was extracted from rumen contents，qRT-PCR was used for the absolute quantification of these related microbes by the consruction of the standard plasmids.The result showed that the number of S.ruminantium on different days of age were significantly different (P＜0.05).However，with the increasing of dietary nutrient levels，the number of P.ruminicola，R.flavefaciens，S.dextrinisolvens，B.fibrisolvens and Methanogens increased significantly (P＜0.05).With the distinct days of age and dietary nutrient levels，pH was not significantly different (P＞0.05).The content of acetate acid showed significantly downward trend from lower dietary nutrient levels to higher dietary nutrient levels(P＜0.05).The ratio of butyrate，isobutyric，pentanoic acid and isovaleric to total volatile fatty acid (TVFA) showed significantly upward trends(P＜0.05).There were mutually improved relationships between different rumen microbes.Propionic acid content was significantly and positively correlated with the number of V.parvula(r＝0.42，P＜0.05)，while the content of pentanoic acid was extremely significantly negatively related to the number of V.parvula (r＝－0.53，P＜0.01).Compared with the effect of days of age，different nutrient levels have greater effects on rumen microbes and VFA levels.There are correlations between the levels of rumen VFA，pH and related ruminal microbes.

 This study was conducted to construct a novel bacterial artificial chromosome (BAC) of herpevirus of turkey (HVT) as a technical platform for generation of recombinant HVT live vectored vaccine.The mini-F sequences were inserted into the genome of HVT in lieu of glycoprotein C (gC) gene through homologous recombination.The mini-F recombinant HVT were selected by GFP and gpt labeling.Then the DNA of mini-F recombinant HVT was transferred into E.coli DH10B cells to construct the HVT BAC.Following identification of correct construction of HVT whole genome BAC through RFLP method，HVT BAC DNA was transferred into E.coli GS1783 cells for further study after checking again.Then the HVT BAC was transfected into chicken embryo fibroblasts (CEF) for rescuing of virus.And also the gC recovered virus was generated by replacement of mini-F sequences with gC gene through homologous recombination again.Finally，the growth characteristics and protective efficacy against Marek’s disease of the gC recovered HVT，mini-F recombinant HVT and the parental virus (HVT FC126) were investigated.Five strains of mini-F recombinant HVT was obtained.One of the five recombinants (HVT^mini-F/ΔgC) was selected for isolation of DNA to transfer into E.coli DH10B cells to construct HVT BAC which was identified successfully through RFLP.Following up，DNA of HVT BAC was transferred into E.coli GS1783 cells successfully to obtain several positive clones and one of these clones was selected and named BAC^HVT-G.The recombinant HVT was rescued successfully from BAC^HVT-G，named HVT^BAC-ΔgC.And the gC recovered virus(HVT^BAC-gC-R) was successfully obtained through homologous recombination.The HVT^BAC-gC-R and HVT FC126 showed no significant difference for growth kinetics or immunity.However，the propagation ability of HVT^BAC-ΔgC was significantly decreased compared to HVT and the immunity against MD of HVT^BAC-ΔgC was also decreased.This study successfully constructed a novel HVT bacterial artificial chromosome，which was an infectious clone of the whole genome of HVT.The gC gene of HVT is a non-essential gene，but plays an important role for propagation of virus.The deletion of gC gene will reduce the immunity of HVT against Marek’s disease.

To study the oral immunogenicity of attenuated Salmonella pullorum，carried HN (haemagglutinin-neuraminidase) gene vaccine of Newcastle disease virus (NDV).In this study，the recombinant plasmid pcDNA3-HN was finally electro-transformed into attenuated Salmonella pullorum ΔcrpC79-13 to construct the recombinant strain ΔcrpC79-13 (pcDNA3-HN).Then the 7-day-old chickens were oral inoculated with 1×10⁹CFU recombinant strain ΔcrpC79-13 (pcDNA3-HN)，and the Δcrp C79-13 (pcDNA3)，PBS，NDV Ⅳvaccine were act as the control group.Then the dynamic level of HI antibody and intestinal mucosal IgA antibody against NDV were determined at 7，14，21，28，35 days post vaccination.At the same time，the level of peripheral blood lymphocyte proliferation of immune chicken and the challenge protection efficiency were determined.The results showed that the recombinant vaccine strains Δcrp C79-13 (pcDNA3-HN) carrying NDV HN gene vaccine was constructed successfully，which could induce HI antibody against NDV at 14-35 d after immunization and reach the highest value at 21 d after immunization.The content of intestinal mucosal IgA antibody in Δcrp (C79-13 pcDNA3-HN) group was slightly higher than that in Δcrp C79-13 (pcDNA3) group，but the difference was not significant (P> 0.05).The peripheral blood lymphocyte stimulation index of Δcrp (C79-13 pcDNA3-HN) group was significantly higher than that of the PBS control group (P<0.01)，and was slightly higher than that of the ΔcrpC79-13 (pCDNA3) group （P＜0.05) at 28 d after immunization.The immunity protection rate against strong virus was 67%（10/15） in experiment group with 10³ EID₅₀ NDV F₄₈E₉，the protection rate was 100% which challenged with 10⁸ CFU Salmonella pullorum C79-13 strain.The results showed that the recombinant attenuated Salmonella pullorum ΔcrpC79-13 (pcDNA3-HN) strains have good oral immunogenicity.

The immune effects of Xinjiang Wild Artemisia rupestris L.crude polysaccharides (WARCP) as an adjuvant and sparing antigen on different doses of influenza virus vaccine (IVV) were explored.ICR mice were intramuscularly immunized respectively with different doses (0.05，0.1，0.5 μg) of the IVV alone or co-administered with 300 μg WARCP at 0 d and 14 d.The growth state of mice were observed，antibody level of IgG and IgG₁，IgG_2a in serum were tested by ELISA；splenocyte proliferations were tested by MTT；The expression levels of CD4⁺IL-4，CD8⁺IFN-γ and CD4⁺CD25⁺Foxp3⁺Treg cells were tested by flow cytometry.The results showed that no significant differences in the body weight were observed between mice from different groups (P>0.05)；WARCP can significantly improve the antibody level of influenza-specific IgG and IgG₁，IgG_2a (P<0.05)；WARCP can significantly promote lymphocyte proliferation，the secretion level of IL-4 and IFN-γ (P<0.05),and significantly reduce the level of expression of Treg cells (P<0.05)；In particularly，it can enhance the humoral and cellular immunity responses of low-dose IVV,indicating an at least 91% reduction in vaccine dosage by adding WARCP as adjuvant.These results indicated that adding WARCP to IVV enhanced the immune efficacy of IVV by reducing the level of Treg cells，significantly promote Th1-type immune response particularly，it can be an adjuvant with the advantages of high safety and dose-sparing.

This study was conducted to investigate the difference in susceptibility to Eimeria tenella infection among different chicken breeds (Gallus gallus domesticus).Twenty-six-day-old，coccidia-free chickens of 24 Chinese indigenous chicken breeds and one commercial breed，recessive white，were divided into two infected groups and one non-infected control group randomly，15 chickens each group.The infected groups were inoculated into crop with 1×10⁵ sporulated E.tenella oocysts per bird.The mortality，relative rate of weight gain，cecal lesion scores，oocyst production and anticoccidial index (ACI) were measured as parameters for evaluation of the susceptibility of different breeds.Among 25 breeds，Langshan，Tibetan，Big bone，Xiaoshan，Wenchang，Anyi tile-like，Buff baier，Xianju，Daweishan mini and Jinhu silky chickens appeared clearly as the least susceptible，with the ACIs ranging from 56.80 to 85.76，showing no death or lower mortality，lower lesion score and higher relative rate of weight gain than other breeds，though higher oocyst production was found in those breeds except Tibetan chicken.Recessive white，Qingyuan partridge，Huiyang bearded，Gushi，Beijing oil，Piao and Wenshang barred chickens were the most susceptible with the ACIs ranging from -11.64 to 18.67，showing higher mortality (except Beijing oil chicken)，lower relative rate of gain，higher lesion scores and higher oocysts production (except Gushi chicken) than other breeds.The rest of the 8 breeds showed moderate susceptibility with the ACIs ranging from 29.81 to 53.36.There are significant differences in the susceptibility to E.tenella infection among different chicken breeds.And the meat type breeds showed higher susceptibility than eggs and meat breeds.

 The cold-inducible RNA-binding protein (CIRP) is an upstream regulator of the NF-κB and ERK pathways，which are essential for the replication of the influenza virus and the initial immune response.In order to investigate the effect of CIRP on the replication of H1N1 influenza A virus and its possible molecular mechanism，in this study，CIRP overexpression BHK-21 (Cirp+BHK-21) cells were constructed，and the phosphorylation levels of NF-κB and ERK in Cirp+BHK-21 cells were detected by Western blot to confirm the effect of CIRP on regulation of NF-κB and ERK1/2；Real-time RT-PCR was used to detect the dynamic changes of virus load in Cirp+BHK-21 and control cells after infected with influenza A virus，the method was also used to detect the dynamic changes of virus load in Cirp+BHK-21 cells which were blocked by the NF-κB inhibitor PDTC.The results of Western blot exhibited that overexpressed CIRP could significantly increase the expression of phosphorylation level of NF-κB (P<0.05)，but had no significant effect on the phosphorylation level of ERK.The results of quantitative detection of virus showed that overexpressed CIRP could significantly enhance the proliferation of influenza A virus，the virus load in the Cirp+BHK-21 cells were 111%，103%，167% and 235% (P<0.05) at 3，9，15 and 21 h PI，respectively，compared to the control group；Blocking the NF-κB was significantly decreased the virus load in the Cirp+BHK-21，and the virus load in treatment group were 98%，42%，19% (P<0.05)，7% (P<0.05) at 3，9，15 and 21 h PI，respectively，compared to the unblock group.Therefore，this study confirmed that overexpressed CIRP could enhance the proliferation of influenza A virus via activation of NF-κB pathway.

The study aimed to prepare oxyclozanide oral suspension and evaluate its physical and chemical properties，and establish an HPLC method for the determination of oxyclozanide.The sedimentation volume ratio and redispersibility were used as variable factors，the single factor experiment and the quadratic regression orthogonal rotational combination design were carried out to optimize the prescription.Then，the shape，size of oxyclozanide suspension was measured by transmission electron microscopylaser and particle size analyzer，and the content of oxyclozanide in suspension was determined by HPLC.The result showed that the suspension (100 mL) was consisted of 3.2 g oxyclozanide，0.2 g carbomer 974p，0.3 g sodium dodecyl sulfate sodium salt，0.02 g methyl 4-hydroxybenzoate and 0.4 g sodium pyrosulfite.With good redispersibility，the sedimentation volume ratio of oxyclozanide suspension was 0.999.Excellent line relationship was obtained in the range of 18-100 μg•mL^-1 (r²=0.999 3).The oxyclozanide suspension was successfully prepared with simple preparation process，good stability and well quality-controlled product.

The aim of the present study was to investigate effects of Fusarium toxins on distribution and mRNA expression levels of intestinal IL-1β and IL-6 in post-weaning piglets.A total of 40 healthy piglets (Duroc×Landrace×Large White) aged at 35 d with an average body weight (8.45±0.94) kg were randomly allocated into two treatments with 20 in the control group (Control) and 20 in the test group (Fusarium Toxins).Piglets in the control group were fed a basal diet only，and piglets in the Fusarium toxins group were fed a test diet (0.90 mg•kg^-1 ZEN，1.43 mg•kg^-1 DON，5.85 mg•kg^-1 FUM).The experimental period was 35 d after 7 d adaptation.Then 10 piglets of each treatment were chosen to slaughter and sample.Results showed because of the Fusarium toxins，the positive cells of the small intestinal IL-1β and IL-6 were from dispersed in the lamina propria to villous lymphocytes concentration.But the IL-1β and IL-6 positive cells of the colon were mainly distributed in the lamina propria cells.And the IL-1β mRNA levels of duodenum，the IL-6 mRNA levels of duodenum and jejunum and the IL-1β and IL-6 mRNA levels of colon of the Fusarium toxins group were significantly higher (P<0.05) than that of the control.The mRNA levels of IL-1β and IL-6 had a correlation between small intestine and colon.In the present study，it suggested that Fusarium toxins exerted a deleterious effect on inflammatory response via distribution and mRNA expression levels of intestinal IL-1β and IL-6，which have a negative impact on immunity of piglets eventually.

The purposes of this study was to explore the expression and location of TGF-β3 and Claudin-11 in the boar testis under heat stress (37-40 ℃) and normal temperature (20-27 ℃).Six boars (Landrace，18 months of age) were used and divided into 2 groups.3 boars were homed in a thermo-controlled temperature (37-40 ℃，3 h daily，consecutive 7 d) house as a heat stress group.After heat treatment，the boars were driven back to normal temperature (20-27 ℃).The other 3 boars were homed in 20-27 ℃ house as a control group.7 days later，all boars were castrated and the testis tissues were harvested.qRT-PCR，Western blotting and immunohistochemistry were used to explore the changes of mRNA and protein in response to heat treatment.qRT-PCR showed that relative expression levels of TGF-β3 mRNA significantly increased （P<0.01），while relative expression levels of Claudin-11 mRNA decreased（P<0.05） in heat treatment group compared with the control.Western blotting found that the expression levels of TGF-β3 protein significantly increased（P<0.05）in heat treatment group，while the expression levels of Claudin-11 protein decreased（P<0.05）compared with the control.Immunohistochemistry results showed that：TGF-β3 immunoreactivity staining was observed in all stages of germ cells and Sertoli cells in both heat stress group and the control，and the depth and area of the positive staining in heat stress group were higher than that of the control；Claudin-11 immunoreactivity staining decreased in heat stress boars compared with the control in that Claudin-11 immunoreactivity staining localized in a consecutive strand area corresponding to the blood-testis barrier in the testis of control boars，while Claudin-11 immunoreactivity staining was limited to Sertoli cells and no obvious immunoreactivity strand that could be found in heat stress group.Then we could get a conclusion that heat stress damaged the sperm quality and spermatogenesis maybe partly via heat stress increased the expression of TGF-β3 and decreased the expression of Claudin-11 in boar testis.

This study aims at establishing a simple method of obtaining the high-purity rat pulmonary microvascular endothelial cells isolated and cultured in vitro.Peripheral lung tissues were separated from 5 to 7 days of SD rat under sterile condition.Rat pulmonary microvascular endothelial cells was acquired by taking the tissue graft block method，digested with trypsin and then maintained at 37 ℃ in a 5% CO₂ humidified atmosphere.The cells were purificated using the method of immune magnetic beads，subsequently identified by the immunofluorescence and flow cytometry，and then determined by MTT colorimetric analysis to detect the original generation of pulmonary endothelial cell growth curve.Rat lung microvascular endothelial cells was successfully obtained by taking the tissue graft block method and individual cells under inverted microscope showed short fusiform or polygon and a typical paying stone appearance with single-layer adherent growth after confluence.The cells were strongly positive for CD31 factor and negative（P=0.1，and P<0.5） for lymphatic endothelial cell specific surface marker VEGFR-3 detected by immunofluorescence staining and flow cytometry respectively.In our study，a method of higher purity of rat lung microvascular endothelial cells culture in vitro was successfully established.