The objective of this study was to clone caprine cationic amino acid transporter gene SLC3A1 and investigate its mRNA expression in different tissues and its development regularity in small intestine.The cDNA sequence of caprine SLC3A1 gene was cloned with the primer which was designed according to mRNA sequence of Ovis aries and Bos taurus in GenBank.Then its expression was quantified by Real-time PCR in 11 tissues from goats at the age of day 1 and duodenum,jejunum and ileum from goats at the age of day 1,month 6,month 8,month 10 and month 12.The results indicated that cDNA sequence of SLC3A1 gene was obtained,and its homology with SLC3A1 gene in Ovis aries,Bos taurus,Sus scrofa,Homo sapiens,Mus musculus and Rattus norvegicus were 99%,97%,88%,86%,80%,79%,respectively.SLC3A1 gene was expressed in all of these collected tissues,whereas the expression level varied from tissues.Specifically,its expression in kidney,jejunum,ileum and colon were significant higher than that in other tissues at the age of day 1 and decreased systematically (P<0.05).The expression of SLC3A1 in small intestine at the same age of goat was ileum>jejunum>duodenum.SLC3A1 gene expression in duodenum at the age of day 1 was significant higher than that at the other ages (P<0.05),it decreased with age in jejunum,and there was no significant difference among ileum with age (P>0.05). In conclusion,SLC3A1 gene and b^0,+ cationic amino acid transporter system were mainly expressed in kidney and intestine.The expression of SLC3A1 gene was differentially regulated and distributed by developmental stages and segments of small intestine in goat.

This experiment was intend to study the changes of long non-coding RNA (H19) expression levels in skeletal muscle development and regeneration,and lay the foundation of its mechanism reach in skeletal muscle development.C2C12 cell line and ICR mice were used as experimental material,bioinformatics assay was used to exploit its non-coding character and low conservatism in different species,and the expressions of H19 in C2C12 cell differentiation,skeletal muscle development and the phase of muscle regeneration were detected by quantitative Real-time PCR (qRT-PCR).The results showed that,H19 expression levels in postnatal mouse skeletal muscle decreased with increasing age;during C2C12 cell differentiation,H19 mRNA increased gradually,and then maintained a high level;The expression of H19 was maintained at a high level through days 4 to 6 after injury.In consideration of its express character in C2C12 cell differentiation and skeletal muscle damage repair model,H19 may play an important role in promoting myogenesis and skeletal muscle regeneration.

The aim of this study was to clone and analyze the expression pattern of buffalo MC1R gene.A pair of specific primers was designed according bovine MC1R sequence (GenBank accession No.:JN123363.1),with genome DNA of swamp buffalo,White swamp buffalo,Murrah buffalo and Yellow cattle as template,MC1R was amplified by PCR.Then relative expression level of MC1R gene of swamp buffalo,White swamp buffalo,Murrah buffalo and Yellow cattle was analyzed using QRT-PCR,and protein expression was detected by Western blotting method.The results showed that the 954 bp coding region of buffalo MC1R gene was successfully cloned and sequenced,which code for 317 amino acids.The MC1R gene nucleotide sequences and amino acid sequences of swamp buffalo,White swamp buffalo,Murrah buffalo and Yellow cattle were highly conserved.Five polymorphic sites were found between White swamp buffalo and swamp buffalo of MC1R gene,including 476 T→C,618 G→C,881 G→A,930 G→A and 931 A→G,which caused three nonsynonymous mutation sites of Phe159Ser,Glu310Ala and Asp294Ala.The QRT-PCR result showed that relative expressions of MC1R gene of Murrah buffalo,swamp buffalo and Yellow cattle were significant higher than that of White swamp buffalo (P<0.05).The Western blotting results revealed that MC1R protein expression level in swamp buffalo was higher than that of White swamp buffalo.In conclusion,there were amino acids mutation in White swamp buffalo MC1R gene,and MC1R gene relative expression of White swamp buffalo was lower than that of other buffalo,which was the main reason of lacking of melanin production in White swamp buffalo.

To construct a lentiviral vector RCASBP carrying the enhanced green fluorescent protein (EGFP) gene which could be expressed stably in DF-1 cell,EGFP gene was amplified by PCR and then inserted into the lentiviral vector RCASBP after digested with the restriction endonuclease ClaⅠto construct recombinant lentiviral vector RCASBP-EGFP.The recombinant vector was transfected into DF-1 cells by Lipofectamine^TM 2000.Avian leukosis virus (ALV) p27 antigen ELISA test was performed after four passages of the transfected cells and the positive results of ELISA suggested the success rescue of the virus.The expression of EGFP was observed in more than 80 percentages of DF-1 cells under fluorescence microscope.The proviral genome PCR showed EGFP gene carried by the recombinant lentiviral vector RCASBP-EGFP had been integrated into the genome of DF-1 cells.The RCASBP lentiviral-mediated expression system provided a basis for study of the structure and function of ALV genes.

To explore the value of Mb1230 protein of Mycobacterium bovis in the diagnosis of bovine tuberculosis,we obtained the Mb1230 gene by PCR and constructed recombinant plasmid pET-22b-Mb1230.Recombinant Mb1230 protein was obtained by IPTG induction and purified by affinity chromatography.The activity of the recombinant protein was evaluated by TST test,IGRA test and indirect ELISA.The size of the recombinant protein matched with the theoretical value proved by SDS-PAGE;Western blotting result showed that the recombinant protein could react with mouse anti-His antibody,and had specific band;The results of TST test,IGRA test and indirect ELISA test also showed the recombinant protein had antigenic activity.The results indicated the recombinant protein Mb1230 had good B cell and T cell activity,so,it had the potential application in the diagnosis of bovine tuberculosis.

In order to further study the mechanism of Lactobacillus acidophilus,GFP labeling shuttle expression vector pSET4s-P1-GFP-P2 was constructed by inserting green fluorescence protein (GFP),upstream and downstream homology arm of Lactobacillus acidophilus based on temperature shuttle expression vector pSET4s.Recombinant Lactobacillus acidophilus expressing green fluorescence gene was constructed by transforming pSET4s-P1-GFP-P2 into Lactobacillus acidophilus using electrotransformation.We could obtain the recombinant Lactobacillus acidophilus by resistance screening and temperature screening,and named it as ΔMG6243(GFP) which could expressed GFP gene stably by fluorescence microscopy observation and Western blotting analysis.We could analyse the genetic stability and biological characteristics with fluorescence microscopy observation and plate count.Green fluorescent of recombinant Lactobacillus acidophilus could be observed in the blue light,and also be observed after 10 generations.There were significant differences neither growth characteristics nor pH and salt tolerance compared with the wild mushrooms.The results showed that GFP labeling recombinant Lactobacillus acidophilus was constructed successfully.Exogenous gene could be non-resistant and integrative to express in the Lactobacillus acidophilus with the vector.It established the foundation for construction of recombinant Lactobacillus acidophilus.At the same time,the role of the GFP as marker laid the foundation for distribution,adhesion of probiotic in animal gastrointestinal tract and the mechanism of Lactobacillus acidophilus.

To establishment a TaqMan Real-time PCR method for detection of duck poxvirus (DPV),we cloned the P4b gene of DPV.The specific primers and probe were designed according to the nucleotide sequence of avipoxvirus available in GenBank.Recombinant plasmid pMD-DPV-P4b was employed as positive standard template for Real-time PCR.By optimization of reaction conditions,a TaqMan Real-time PCR method for detection of DPV was established.The results of specificity test proved this method had no cross-react with other waterfowl vial agents and poxviruses including avian influenza virus,duck flavivirus,duck hepatitis virus,Newcastle disease virus,duck entertitis virus,goose parvovirus,goatpox virus and fowlpox virus.The detection limit of the assay was 1.29×10² copies/μL of viral DNA,which was 100 times higher than that of the routine PCR.Reproducibility test showed that the CVs of intra assay and inter assay were both less than 2%.Above results supported that the assay was suitable for the detection of DPV very well.

The assay was aimed to study the function of Lin28 gene of sheep,in this study,according to the Lin28 gene mRNA sequence of mice,pigs,cattles and human in GenBank,the degenerate primer was designed.Total RNA was extracted by Trizol from intestine of 100 d sheep embro,and CDS sequence of the gene was amplified by RT-PCR using the primer.Using a series of bio-software such as DNAMAN,BioEdit and NCBI BLASTn,PlantsP,ScanProsite online and so on,we analyzed the homologies between nucleotide and deduced amino acid sequences,constructed phylogenetic tree,analyzed protein functional domains and deduced three-dimensional structure of amino acid sequence.The results showed that the sequence of sheep Lin28's CDS including stop codon,was in a length of 630 bp,encoded 209 amino acids.Analysis of homology showed that sheep Lin28 gene also shared 53.8%,55.2%,70.2%,88.3%,88.7%,89.2%,89.5%,90.5%,95.9% and 98.9% nucleotide homologies and 76.5%,67.9%,93.2%,99.3%,97.7%,99.3%,99.3%,98.5%,99.3% and 100.0% amino acid identities with Xenopus laevis,Danio rerio,Gallus gallus,Bos taurus,Equus caballus,Felis catus,Sus scrofa,Homo sapiens,Rattus norvegicus and Mus musculus,respectively.Two zinc finger domains and one cold shock domain could be found by bioinformatics analyzing.The successful cloning of Lin28 revealed that the gene in the structure and function was very conservative and helpful to study gene function and its role in somatic reprogramming in future.

This assay was aimed to express the PuAH gene of Edwardsiella tarda (E.tarda) EIB202 and analyze its partial characteristics.A pair of specific primers was designed based on the PuAH gene sequence of E.tarda published in GenBank(accession No.:CP001135,CDS No.:ETAE_0818).The gene was cloned and expressed by recombinant plasmid pET32a-PuAH in E.coli BL21(DE3).The recombinant protein was purified and used to immune New Zealand rabbits to prepare antisera.Then the characteristics of expression product and antisera were analyzed by ELISA,Western blotting,hemolysis test and bacterial agglutination test.In conclusion,the expression product was fusion protein with 42 ku molecular weight,and had immunogenicity and antigenicity,but no hemolytic to erythrocytes of eel.Yet the antisera could inhibited the hemolytic of outer membrane protein of E.tarda EIB202 to some extent,and could agglutinate E.tarda EIB202.

Duck Tembusu virus (DTMUV) was newly emerged which caused severe egg-drop syndrome in duck.Here,we constructed the recombinant baculovirus by honeybee melittin signal peptide (Mels) which expressed E protein of DTMUV JSXZ strain,and its immunogenicity and protective efficacy were investigated in duck infection model.Western blotting and indirect immunofluorescence assay (IFA) showed E protein was effectively secretory expressed in supernatant of Sf9 insect cells.Indirect ELISA result showed E protein could induce high levels of anti-E IgG in the sera.MTT result showed that E protein could stimulate the proliferation of T lymphocytes.Moreover,the protection rates were 80% in 0.3 mL group and 100% in both 0.6 mL and 0.8 mL groups,respectively,whereas,all ducks in PBS control group showed the typical clinical signs with 3 died post challenge.These results laid a foundation for the study of DTMUV subunit vaccine.

A strain of virus was isolated and identified from feces of a suspected canine parvovirus (CPV) infected dog.It was eventually identified as CPV using electron microscope examination,hemagglutination (HA) test,animal regression experiment and molecular biology idenfication,designated as BJ-24 strain.The VP2 gene sequence analysis of BJ-24 strain showed that BJ-24 strain belonged to CPV-2a subtype.The analysis of VP2 gene indicated that the BJ-24 VP2 gene shared higher homology of nucleotide with 19 CPV strains available in GenBank and were more than 98.7%.The highest homology was 100.0% with CPV-JS2 strain.Phylogenetic tree analysis showed that BJ-24 strain belonged to China cluster and there was the closest distant between BJ-24 strain and CPV-JS2 strain.This study provided the basic evidence for further study on molecular epidemiology in Beijing and laid the solid foundation for developing vaccine and preventing the spread of CPV.

Glutamate dehydrogenase (GDH) which exists in 35 serotypes of Streptococcus suis,is an immunogenic protective antigen.Therefore,the GDH gene was amplified by PCR method,and the GDH gene PCR fragment was inserted oriented to pET-32a to construct a new recombinant expression vector pET-32a-GDH.The E.coli BL21(DE3)(pET-32a-GDH) was cultured and induced to express the rGDH protein.New Zealand White rabbits were immunized by the purified rGDH protein for immunity protective test to determine the immunogenicity of the protein.The results showed that the recombinant expression vector pET-32a-GDH was successfully constructed,and the rGDH protein was efficiently and solubly expressed.70% of model animals immuned by the rGDH protein with propolis as adjuvant were protected after injection of Streptococcus suis type 2 as a virulent pathogen.The experiment results provided technical support for the development of subunit vaccine of Streptococcus suis.

In order to enrich the biological characteristics of VP gene from Muscovy duck parvovirus Zhejiang isolate (MDPV-ZJ),the target VP gene fragments were amplified by PCR method with specific primers,then the obtained PCR products were cloned and sequenced.The bioinformatics analysis of VP gene of MDPV-ZJ was conducted.The results revealed that MDPV-ZJ VP gene was 2 199 bp in length,coding an open reading frame (ORF) with 732 amino acids.The molecular weight,theoretical isoelectric point,instability index and grand average of hydropathicity of MDPV-ZJ VP gene were 81.32 ku,6.59,37.49 and －0.667,respectively,and with no signal peptide.The genetic evdution ary tree based on the VP gene showed that the MDPV isoaltes contains two groups:The typical MDPV and recombinant MDPV; Also,the typical MDPV could be divided into three branches:Taiwanese branch,Maniland China branch and Euro branch,which existed obvious regional genetic evolution relationship.In this assay MDPV-ZJ was belonged to MDPV Maniland China branch.

This study was aimed to prepare excellent fluorescence immunochromatographic strip for clenbuterol (CLB) which could achieve the purpose of rapid and quantitative detection and laid a foundation for its industrialization.The strip was prepared through single factor experiments using latex particles as a fluorescent label.The preparation process was optimized with CLB concentration,drying time,mixing reaction time,reaction temperature and chromatography time.And the properties of this strip including detection limit,specificity,clinical accuracy and added recovery,degree of precision and stability were investigated.Optimum modification conditions were shown as following:CLB concentration was 2.0 mg/mL,drying time was 5 d,mixing reaction time was 1 min,reaction temperature was 25 ℃,chromatography time was 15 min.The detection limit was up to 0.35 μg/L and the cross rate between CLB and other eight similar drugs were less than 0.8% which demonstrated the good specificity.The added recoveries of each batch were between 80% and 120% which met the requirements.The coefficient of variation (CV) under different concentrations of CLB was less than 5% which met the precision standard.Besides,the stability of the experiment also showed that the test strip had good stability.

The aim of this study was to establish a multiplex PCR method for simultaneously detecting porcine reproductive and respiratory syndrome virus (PRRSV),porcine circovirus type 2 (PCV2),porcine pseudorabies virus (PRV) and Mycoplasma hyopneumoniae (Mh).Four pairs of primers were synthesized according to the reference.The multiplex PCR method was developed by optimizing the reaction condition,specificity and sensitivity detection.Four different amplicons with size of 424,490,298 and 360 bp for PRRSV,PCV2,PRV and Mh,respectively,were yielded.The sensitivity of multiplex PCR indicated that the detection limit was 3 copies by using Multiplex PCR Master Mix,and other common pathogens were not amplified.A total of 60 specimens from piglets were tested by multiplex PCR method.The positive accordance rate between simple and multiplex PCR was 100%.This study indicated that multiplex PCR might be a useful tool for rapid and sensitive etiological diagnosis and provided an effective technical support for pathogenic molecular epidemiology investigation.

In 2013,one case of suspected H9 subtype avian influenza occurred in a chicken farm of Jilin povince.Clinical samples were collected from the diseased farm,inoculated into the allantoic cavity of 9-day-old SPF chicken embryo,and then one strain of virus was isolated.The results of HA test,HI test and molecular biology test all showed that the isolate belonged to H9 subtype avian influenza virus (AIV).The HA cleavage site of the isolate was RSSR↓GLF,which was consisted with the molecular characteristic of low pathogenic AIV.The HA peptide chain had 9 potential glycosylation sites which were same as other isolates of recent years.The isolate had 8 receptor binding sites,including 234 receptor binding site by glutamine (Q) mutating into threonine (T).The phylogenetic tree revealed the isolate belonged to Eurasian lineages and it had far genetic relationship with the earliest domestic isolate (A/Chicken/Beijing/1/94(H9N2)),but had close genetic relationship with the representative strain (A/Chicken/Guangxi/55/2005(H9N2)) of major epidemic branch since 2007.We prepared an inactivated oil-emulsion vaccine of the isolate,and then vaccinated SPF chicken.21 days after vaccination,the HI titer of chicken serum antibody reached up to 10log2.The result suggested the isolate had good immunogenicity.

An indirect competitive enzyme-linked immunosorbent assay (ELISA) for detecting tilmicosin residue in milk was developed and applied to milk samples after the determination of dilution of coating antigen and antibody to timicosin by using checkerboard method,and the optimization of coating condition,reaction time and temperature,and reaction time of enzyme-labled secondary antibody.The results showed that the 50% inhibition concentration (IC₅₀) of the indirect competitive ELISA was 2.1 ng/mL and the limit of detection (LOD) of tilmicosin in milk was 2 μg/L.The recoveries of tilmicosin added in milk at 5,10 and 20 μg/L ranged from 79.2% to 90.1% with the coefficients of variation (CV) not higher than 9.0%.The indirect competitive ELISA would not react with other common macrolide drugs.The sensitivity of indirect competitive ELISA developed in the study complied to the maximum residue limit of tilmicosin set by China,and laid the foundation for the test kit of the rapid detection of tilmicosin in milk.

In order to evaluate whether Haemophilus parasuis espP gene could use as a vaccine candidate antigen or not,the espP gene was amplified by PCR using the specific primers that designed according to corresponding sequence getting form GenBank (accession No.:HAPS_1381) and then sent to sequence.After translating the DNA sequence into amino acid sequence,the signal peptide,hydrophilic region,linear B cell epitopes and 3D structure of Hps espP protein were predicted using multiple bioinformatics analysis.The results showed that the full length of Hps espP gene was 2 343 bp,encoding 781 aa.The first 23 aa were predicted as signal peptide.There were 12 hydrophilic regions and 12 B cell epitopes in the Hps espP protein.The 3D structure of C-terminal of the protein was consisted of a monomeric β-barrel containing 12 β-strands and a central pore.The study laid the foundation for further development of molecular vaccines based on espP protein against Hps infection.

Peroxisome proliferators-activated receptor-γ (PPAR-γ) coactivator-1α (PGC-1α) is a versatile transcriptional regulator.PGC-1α is characteristically involved in the regulation of several crucial cellular pathways including mitochondrial biogenesis,fatty acid oxidation,gluconeogenesis and so on.PGC-1α is significantly associated with meat quality,since it may affect the fiber-type switching through several signaling pathways in livestock and poultry.Therefore,study on PGC-1α expression regulation in the transcription and post-translation level can provide new strategies for improving the meat quality.In recent years,the effect of dietary nutrition on PGC-1α has been also researched gradually for its importance in the regulation on meat quality in livestock and poultry.This review described the structure and expression patterns in tissues of PGC-1α,and its regulatory mechanism on muscle fiber-type switching,focusing on recent advances on its expression regulation in the transcription and post-translation level,also summarized the effect of dietary nutrition on PGC-1α.

Myogenic regulatory factors (MRFs) are important genes controlling myogenesis,including MyoD,MyoG,Myf5 and Myf6.MyoD is able to convert nonmyogenic fibroblast cells into myogenic cells,being a master regulatory gene in the process of muscle specified transcription;MyoG can control the start of myoblasts fusion,and induce myoblasts proliferation;Myf5 works in coordination with MyoD in regulating myogenesis,and Myf6 cooperates with MyoG to control myoblast determination.Studies indicate that the expression profiles of MRFs are closely related to meat production and meat quality in meat-type livestock,and the gene activity can be significantly affected by genetic,nutrition and environment.This article reviewed that the structure,function,expression profile and activity regulation of MRFs and the extrinsic regulation functions of nutrition,light,temperature,exercise and active factors on the gene activity of MRFs and myogenesis,aiming to provide the theory basis for manipulating the muscle development of animals.

This experiment was conducted to study the effect of amino acid balance diet on production performance,serum biochemical indexes and economic benefits in dariy cows.36 healthy Holstein cows in the early lactation period were selected,the average number of lactation (DIM) was (39.69±12.48) d,the average parity was 3.02±0.64.Principle of experiments using single factor randomized experimental design accroding to age,milk yield,parity identical same or similar,the 36 Holstein cows were assigned by random block design to 3 groups,and each group were 12 cows.The control group cows were fed with TMR diets only adding rumen-protected methionine (HMBi);Diets of group 1 and 2 were on the basis of control group,respectively adding two kinds of rumen-protected lysine levels in order to balance the amino acid in diet.The results showed as follows:① Compared with control group,the feeding conversion ratio in groups 1 and 2 were increased by 10.60% and 11.26% (P>0.05),respectively,while the dry matter intake had no significantly changed (P>0.05),the 4% average daily milk yield in groups 1 and 2 were significantly increased by 8.65% and 10.58% (P<0.05),respectively,and the milk fat and milk protein yield in groups 1 and 2 were significantly or extremely significantly increased (P<0.05;P<0.01),respectively.Adding amino acid balance diet had no significantly effects on reproductive performance and health conditions in dariy cows (P>0.05).② When feeding the amino acid balance diets for 60 days,the contents of GLU and TAA in serum of dairy cows were significantly increased (P<0.05),while the BUN content was significantly decreased (P<0.05).③ Compared with control group,the economic benefits of groups 1 and 2 were increased by 15.09 and 16.36 yuan/(head·d).The results indicated that feeding the amino acid balance diets could significantly improve production performance of dairy cows,and there was a tendency to improve metabolic conditions.

5 male Small-tail Han sheep with the body weight of 45 kg at 1.5 years old,fitted with permanent rumen fistula,to study of the effect of the supplement of the untreated commercial lysine on the rumen microbiocoenosis,rumen metabolism and whole body metabolism by 5×5 Latin square design (5 groups:control,control with low urea (equal nitrogen),low lysine,control with high urea (equal nitrogen),high lysine).The amounts of lysine-HCl supplement were 0,4.0 and 8.0 g/kg diet (as DM basis) in each group respectively,with two urea supplement control groups of equal amount of nitrogen.The results showed that by supplement of 4.0 and 8.0 g/kg of lysine-HCl diet (as DM basis) respectively,the voluntary intake of dry matter of sheep were increased by 5.5%(P>0.05) and 11.8%(P<0.05) respectively.The total amount of rumen bacteria were increased by 18.9%(P<0.01) and 23.9%(P<0.01) respectively,as the amount of rumen Coccus was increased by 21.2%(P<0.01) and 30.1%(P<0.01) respectively.The amount of rumen big Bacillus was decreased by 16.7%(P<0.05) and 33.3%(P<0.01) respectively.The total VFA of was increased by 9.6%(P<0.05) and 12.2%(P<0.01) respectively,as the butyric acid was increased by 12.8%(P>0.05) and 20.2%(P<0.01) respectively.The apparent digestibility of organic matter was increased by 6.4%(P>0.05) and 10.0%(P<0.05) respectively,the apparent digestibility of crude protein was increased by 7.0%(P>0.05) and 18.3%(P<0.01) respectively;And nitrogen retention was increased by 46.4%(P>0.05) and 110.7%(P<0.01) respectively.However,there was no effect of lysine supplement on the amounts of the rumen protozoa and fungi,and the concentration of rumen ammonia nitrogen.It was concluded that the supplement of untreated commercial lysine to adult sheep would increase the voluntary intake,digestibility,nitrogen retention the rumen microbiocoenosis and the total amount of rumen bacteria,especially the rumen Coccus of adult sheep.The effect of lysine supplement could not be replaced by urea supplement completely.

This experiment was conducted to investigate the effect of soybean meal replaced by brewer's grain (BSG) or fermented brewer's grain (FBSG) on growth performance,immune organ index and intestinal flora of meat geese.A total of 250 geese at 28 days of age were randomly assigned to 5 groups with 5 replicates per group and 10 geese per replicate.The geese in control group were fed a basal diet (with soybean);The geese in groups 1 and 2 were fed the basal diet with the 10% and 20% BSG replacing the soybean,respectively;The geese in groups 3 and 4 were fed the basal diet with the 10% and 20% FBSG replacing the soybean,respectively.The whole experiment period was 42 days,including three phases:15 d (early stage),14 d (middle stage) and 14 d (later stage).The results showed as follows:①From the whole experiment period,compared with control group,the ADG of geese in group 1 was significantly increased (P<0.05),the ADG and ADFI of geese in group 4 were significantly decreased (P<0.05),there were no obvious change in growth performance of geese in groups 2 and 3 (P>0.05).② Compared with control group,the thymus index and spleen index of geese in groups 1,2 and 3 were extremely significantly increased at early stage (P<0.01);At middle and later stages,the spleen index of geese in group 2 were significantly and extremely significantly increased (P<0.05;P<0.01);There were no significant differences in bursa of fabricius index of geese among all groups (P>0.05).③ Compared with control group,the number of Escherichia coli and Salmonella in cecum in groups 1 and 2 were extremely significantly decreased at early and later stages (P<0.01),while the number of Bifidobacteria and Lactobacillus were increased in different degrees;the number of Bifidobacteria and Lactobacillus in cecum in groups 3 and 4 were extremely significantly increased at later stage (P<0.01).In conclusion,10% BSG replacing soybean in the basal diet could improve the growth performance and intestinal flora of meat geese;10% FBSG replacing soybean in the basal diet could promote the growth of beneficial bacteria of meat geese,but had no effect on growth performance;While 20% replacing soybean in the basal diet could decrease the feed intake and weight gain of meat geese,hinder its growth and development.

This experiment was conducted to study the effect of soybean meal replacement by fermented rapeseed meal on growth performance,meat quality and serum biochemical indexes of broilers.A total 400 23-day-old Yellow-feathered male broilers were randomly divided into 4 groups with 4 replicates per group and 25 broilers per replicate.The broilers in control group were fed a basal diet,and the others were fed the basal diets with the 3% (group Ⅰ),6% (group Ⅱ) and 9% (group Ⅲ) fermented rapeseed meal equal-nutritionally replacing the soybean meal,respectively.The experiment lasted for 43 days.The results showed as follows:① Compared with control group,the ADG,ADFI and F/G of groups Ⅰ,Ⅱ and Ⅲ showed no significant differences (P>0.05);The ADG of group Ⅲ were 16.47% (P<0.05),15.03% (P<0.05) higher than that of groups Ⅰ and Ⅱ,the F/G of group Ⅲ were 7.71% (P<0.05),4.27% (P>0.05) lower than that of groups Ⅰ and Ⅱ.② There were no significant differences in the pH₁,pH₂₄,meat color (L^*,a^*,b^*),cooking loss and tenderness of chest muscle among all groups(P>0.05);Compared with control group,the water loss rate of groups Ⅰ,Ⅱ and Ⅲ were significantly decreased (P<0.05).③ Compared with control group,the serum GLU content of groups Ⅰ,Ⅱ and Ⅲ were decreased by 13.06%(P<0.05),8.12%(P>0.05) and 9.57%(P>0.05);The serum TP content of groups Ⅰ,Ⅱ and Ⅲ were increased by 2.50%(P>0.05),20.86%(P<0.05) and 33.92%(P<0.05);The serum GPT content of groups Ⅰ,Ⅱ and Ⅲ were decreased by 7.99%(P>0.05),18.85%(P>0.05) and 26.98%(P<0.05).It was concluded that it would be feasible to replace the soybean meal with 3% to 9% fermented rapeseed meal in broiler feeding,and the optimum supplemental level of fermented rapeseed meal was 9%.

This study was designed to investigate the effects of dietary type and crude protein level on production performance and carcass traits of Sichuan mountain Black-bone chicken.The experimental design was a 2×2 factorial arrangement with two dietary types (corn-soybean meal and corn-soybean meal-yeast protein) and two crude protein levels (16.5% and 14.5%).Four hundred and eighty 23-week-old Sichuan mountain Black-bone chicken were randomly divided into 4 groups with 4 replicates per group and 30 chickens in each replicate.The pre-test period lasted for 14 days and the trial period lasted for 120 days.The results showed that the production performance of Sichuan mountain Black-bone chicken was not significantly affected by dietary type (P>0.05),while the average egg weight of dietary crude protein level 16.5% was significantly higher than that of dietary crude protein level 14.5% (P<0.05).It was also demonstrated the average egg weight was significantly affected by the interaction of dietary type and crude protein level (P<0.05),but laying rate,daily egg production,daily feed intake and feed/egg were not significantly affected (P>0.05).However,the carcass traits of Sichuan mountain Black-bone chicken was not significantly affected by dietary type and crude protein level or the interaction of the two factors (P>0.05),although dressing percentage,percentage of eviscerated yield,percentage of half-eviscerated yield,percentage of breast muscle and percentage of leg muscle of the dietary type of corn-soybean meal-yeast protein (crude protein level 16.5%) displayed better value.These results indicated that the average egg weight of Sichuan mountain Black-bone chicken was affected by dietary crude protein level greatly.The dietary type of corn-soybean meal-yeast protein (crude protein level 16.5%) was better for Sichuan mountain Black-bone chicken by comprehensive evaluation of production performance and carcass traits.

Medicago sativa is one kind of excellent legumes and flavonoids are secondary plant substances which widely distribute in the natural world.This ingredient has bioactivity such as estrogenic effect,antioxidant and cardiovascular system improvement.The article reviewed the flavonoids bioactivity of Medicago sativa on animal and provided reference for further research and development of flavonoids of Medicago sativa.

The genetic polymorphisms in PLIN gene were detected by PCR-RFLP method in four Chinese local chickens including Jining Bairi chicken,Laiwu Black chicken et al.,and one breeding strain.The correlations between the SNP and the carcass and fatness traits were analyzed.As a result,a novel G→A mutation at 2 467 bp in PLIN gene was identified in the five populations.Three genotypes (A1A1,A1A2 and A2A2) were detected,and allele A1 was predominant in all the five experimental populations.The statistical analysis showed that the 2 467 bp polymorphism locus was significant association with some carcass and fatness traits (P<0.05).The living body weight,carcass weight,eviscerated weight,abdominal fat weight and percentage of abdominal fat of A1A1 genotype was higher than A2A2 genotype in chickens (P<0.05).The breast intramuscular fat of A1A2 genotype was higher than that of A1A1 and A2A2 genotypes in chickens (P<0.05).The results showed that PLIN gene had effect on carcass and fatness traits in chickens.

Newborn ovary homeobox gene (NOBOX) is one of maternal genes,which is important in early oogenesis.In order to know the expression pattern of NOBOX gene in porcine tissues,Real-time fluorescence quantitative PCR was used to detected the expression of NOBOX gene in porcine tissues,and then the probe of NOBOX gene was generated and in-situ hybridization were used to detect NOBOX gene localization in porcine ovary.The results showed that NOBOX gene expression was significantly higher in ovary than in other tissues (P<0.01).There was no significant difference in NOBOX gene expression among kidney,liver,mammary gland,muscle and adrenal (P>0.05).In ovary,NOBOX gene was localized in oocyte cytoplasm specifically,but not in granulosa cell.All these results revealed that NOBOX gene was highly expressed in the cytoplasm of procine oocyte,indicating that it played an important role in porcine oocyte and early embryo development.

In this study,the statistical analysis data of 185 340 Holstein dairy cows daily yield records were collected from 2011 to 2014 years in Changji area of Xinjiang,five kinds of models (Wood,IQP,Wilmink,ML,AS models) were used for the first,second,third,fourth and all parities fitting lactation curve.The results showed that the Wood,IQP,Wilmink,ML,AS models fitting precision were within the range from 0.9562 to 0.9828,0.9467 to 0.9809,0.8671 to 0.9752,0.8752 to 0.9175,0.8775 to 0.9127;Milmink model was the best fitting model for the first parity lactation curve,Wood model was the best fitting model for the second,third,and fourth parities lactation curve.

The paper aimed to analyze and validate the function of differentially expressed ENO1 in the cow with retained foetal membrane.In our research,we chose three healthy Holstein dairy cows and three Holstein dairy cows with retained foetal membrane of similar age,foetal times,weight and milk yield and divided them into two groups.The total protein of maternal placenta was extracted in the control and retained foetal membrane of cow.The differential expression of proteins were found out by the 2-DIGE,and the differential expression of ENO1 was validated by the Western blotting and Real-time quantitative PCR.The results showed that ENO1 was significant expression and large multiple in the two groups,and it was significant increased expression in the retained foetal membrane of cow after Western blotting and Real-time quantitative PCR (P=0.015<0.05;P=0.001<0.01).The ENO1 participated in the energy homeostasis,immune and fibrinolysis process which related to the retained foetal membrane of cow.It suggested that ENO1 was likely connected with the retained foetal membrane.

This study was designed to study the possibility of gene drift of the antibiotic marker gene neo in transgenic pigs.First,dominance and recessiveness of neo gene in 6 piglets of F1 were identified using Southern blotting.It was found that 3 piglets were positive transgenic piglets and the other 3 piglets were non-transgenic piglets.The neo gene in blood and digestive tract tissue of experimental piglets were detected using PCR.The result showed that no gene drift was showed in blood and digestive tract tissues of experimental piglets.Then PCR technology was used to detect neo gene in intestinal bacteria of experimental piglets.The result showed that neo gene did not exist in the genome of intestinal bacteria.This study suggested that no gene drift was found in blood,intestinal bacteria and digestive tract tissues of piglets.

The aim of this study was to investigate the developmental patterns of ApoCⅡ gene mRNA in liver in Mashen and Large White pigs, and study the relationship between the expression level of ApoCⅡ and the lipid metabolism in pigs.The mRNA relative expressions of ApoCⅡ gene in liver at seven stages of 1,30,60,90,120,150,and 180-day old in Mashen and Large White pigs were determined by quantitative Real-time PCR.The results showed that the developmental trend of ApoCⅡ mRNA expression in liver between Mashen and Large White pigs was different.The ApoCⅡ mRNA abundance was decreased from birth to 60-day old,then increased at 90-day old,and decreased again after that in Mashen pig.However,the relative expression amount in Large White pig was gradually decreased from birth to 150-day old and increased again at 180-day old.Except for the ApoCⅡ mRNA expression amount at 1-day old,the differences of the expression amount at other stages in Mashen and Large White pigs were significant or extremely significant (P<0.05 or P<0.01).The ApoCⅡ mRNA expression in liver was affected by age and breed,and could play an important role in lipid metabolism in pigs.

This review focused on germplasm resources of domestic sika deer,the genetic background was discussed,characteristics and performance of improved varieties based on Cervus nippon hortulorum Swinhoe were compared and biological basis,genetic trait and performance of crossbreed derived from interspecific cross between domestic sika deer and wapiti were analyzed.For the present status of domestic sika deer,the conservation and utilization methods were provided.

Bovine somatic cell nuclear transfer (SCNT) is a sophisticated technique system,including oocyte maturation,donor cell preparation and oocytes microscopic operation,fusion,activation and culture.Although the birth of cloning cattle has been reported recently,the efficiency of somatic cell cloning has remained lowly.In order to establish the optimization somatic cell nuclear transfer system of Yanbian Yellow cattle,this review summarized only from 6 main aspects mentioned above in this field.

The latest research progress on domestic and international rabbit genetics and breeding in 2014 were reviewed in the present paper based on three aspects of traditional breeding,molecular breeding and reproductive technology,so as to provide references for rabbit breeders and researchers.At abroad,the researchers focused on the analysis of productive performance of different meat rabbit breeds in aspect of traditional breeding,on fur color and productive performance in molecular breeding,meantime rabbit genomic structures were studied by high-throughput sequencing technology.While the key point was still efficient reproductive technique in foreign breeding studies,including semen cryoprotectants development and embryo cryopreservation technology.In domestic rabbit genetic and breeding,molecular bases were still the point of importance for traits of fur,growth and carcass.Secondly,traditional breeding mainly contained evaluation and comparison of Chinese local breeds,cultivating breeds and imported breeds,meantime rabbit cross-strains and modern breeding software development have become two spot light of internal genetic and breeding field in 2014.While studies on rabbit reproduction in domestic rabbit fields were less than these of foreign countries and disperse.Overall,the genetic and breeding studies focus on partial different aspects at home and abroad.For our country,researches on efficient reproductive technology and molecular bases of productive traits should be given further emphasis.

In February 2014,piglets of a certain 100 sows,fed in a large-scale pig farm which had 2 000 sows,died acutely and densely in one week with vomit,emaciation and lethal watery diarrhea after sudden snowy weather.By field survey,pathological anatomy,histopathological observation,immunohistochemistry,reverse transcription-polymerase chain reaction (RT-PCR),sequencing and enzyme-linked immune sorbent assay (ELISA),porcine epidemic diarrhea virus (PEDV),co-infecting with porcine circovirus 2 (PCV2),was confirmed to be the major agent.By adopting measures,including emergency vaccination of sows,egg yolk antibody oral inoculation of piglets,and insulation and disinfection,the disease was successfully and speedily controlled.

To understand the overall status and focus of bovine mastitis vaccine research,knowledge maps of mainly country,institution,author,cite journal,cite reference,keyword,year distribution of publications on research of bovine mastitis vaccine were studied by Citespace Ⅲ,which collected 4 205 international scientific literatures download from Web of Science^TM core collection (1995 to 2015).The results showed that in research field of bovine mastitis vaccine,the number of published papers increased from 1995 to 2012,but it began to decrease in 2013;Countries and institution involved in the research were mainly distributed in America,Europe and Asia,and universities played important roles in this field;In addition,the research field were mainly focus on:Isolation and identification of pathogens,detection of drug susceptibility and pathogenesis exploration;Immune response and relevant regulatory factor in infected dairy cows;Best period for prevention and treatment;Screening for candidate antigen.In future time,studies will pay more attention on research of NF-κB and biofilm in field of bovine mastitis vaccine.

To understand the pathogenic causes of weaning diarrhea in Yunnan province,we collected 210 feces and 60 drinking water,a total of 270 samples in 30 farms with weaning diarrhea and analyzed the virus and pathogenic bacteria.The results of the study showed that drinking water was sever polluted by pathogenic Escherichia coli,40 strains of pathogenic Escherichia coli were detected in drinking water,the positive rate was 66.7%,and 100 strains in feces with positive rate of 47.6%;10 strains of pathogenic Salmonella were detected in drinking water,the positive rate was 16.7%,and 24 strains in feces with positive rate of 11.4%,and were sensitive to imipenem.PCR amplified positive rates of porcine epidemic diarrhea virus (PEDV),porcine circovirous type 2 (PCV2),pseudorabies virus (PRV),classical swine fever virus (CSFV),porcine reproductive and respiratory syndrome virus (PRRSV) and porcine rotavirus (PoRV) in feces were 5.2% to 13.0%,and they were not detected in drinking water.By analyzing the causes of diarrhea in Yunnan province in weaned piglets analysis,understanding the specific causes of diarrhea in weaned piglets,the results provided a more reasonable and more realistic theoretical basis and experimental reference for prevention and control of the disease from the cause.

In order to study the main reason of bacterial goat abortion in Guizhou province,and analyze the bacteria impact of abortion goat immune balance in normal pregnancy animals,107 samples of the goat fetus,uterus and vaginal swabs were collected from clinical health and morbidity abortion goat in 12 scale goat farms of 6 areas in Guizhou province.The bacteria were isolated and identified.The drug susceptibility disc agar diffusion method was launched for the survey of the resistance of three main sources of goat isolates to 20 kinds of antibiotics.The BALB/c mice model of abortion was established using abortion bacteria isolated from goats.The pregnancy related hormones and cytokines were determined by ELISA method.The results showed that 302 strains bacteria of 13 genera were isolated from the samples,and the species of bacteria isolated from morbidity abortion goat were more than the clinical health goats.The bacteria resistant rate of goat reproductive organs in abortion goats was higher than that of clinical healthy goats bacteria.The varying degrees of abortion of BALB/c mice could cause by Escherichia,Staphylococcus and Streptococcus isolated from goats,the rates were Escherichia>Staphylococcus>Streptococcus,respectively and the embryo resorption rates were Escherichia>Staphylococcus>Streptococcus,respectively.Compared with the control group,IFN-γ and IL-2 in the serum were increased;However,IL-4,IL-10 and progesterone were decreased.The results indicated that the occurrence of bacterial goat abortion diseases had certainly related to the number of bacterial types and the level of drug resistance in goat reproductive organs.The levels of pregnancy related hormones and cytokines in goats serum could be changed by abortion bacteria,thus lead to goats abortion.

In order to provide therapeutical guidance for drug admistration,the bacteria from five dead minks suffering with typical symptoms provided by mink farms in Shandong province were isolated and identified,and the drug sensitivity was tested.The bacteria were isolated with TSA plates,and identified using biochemical methods and PCR assay.The drug sensitivity of the isolates to antimicrobial agents was investigated using K-B method.PCR was used to detect the resistance genes.A total of 5 Pseudomonas aeruginosa isolates were obtained from sick minks.The drug sensitivity results showed that 5 strains were sensitive to fluoroquinolone,the third and fourth generations cephalosporin drugs,but were resistant to aminoglycoside,tetracycline,chloramphenicol,penicillin,the first and second generations cephalosporin drugs.There were six resistance genes were detected,aadA1,aac(3')-Ⅱc,aac(6')-Ⅰb,tetA,tetK and cat2.All of the isolates were detected more than three kinds of resistance genes.The result showed that 5 strains were all Pseudomonas aeruginosa,and Pseudomonas aeruginosa was the main cause of mink hemorrhagic pneumonia.The resistance of 5 strains were very serious and mainly for multiple drug resistance phenomenon.The resistance genes detected in the mink were various,and could cause the resistance to the drugs.

In order to isolate the strains which could degrade fiber from the straw,the straw samples were inoculated on the potato agar medium and cultured at room temperature.The isolating strains were inoculated on the cellulose-congo red agar medium,the strains were screened that could produce bigger cellulose-decomposing zone.It was identified by morphological characteristics and molecular biological methods,and characterization of cellulase was analyzed preliminarily.The results showed that the fungus was Aspergillus niger,it could grow at room temperature,and the H/C was 7.64.The highest cellulase activity reached 42.18 U/mL when cultivated at 20 ℃ for 5 days.The optimum pH was 7.0 and the optimum reaction temperature was 20 ℃,the relative cellulase activity still retained above 90% at 20 to 40 ℃ or pH 6.0 to 8.0 for 1 h.The Aspergillus niger was a cold-adapted cellulaseproducing strain,it had strongger producing cellulase ability,and was tremendous potential valuable for microbial development.

The study was aimed to rapidly diagnose two ill and dead cattle in three bay town of Yanji city in Jilin province.We collected liver,spleen,heart,muscle and other tissues to diagnose Clostridium chauvoei infection by epidemiological investigation,clinical autopsy,microscopic obsevation,bacterial culture observation,biochemical experiment,molecular biology diagnosis and animal experiment.The muscles of the ill and dead cattle contacted with crepitus,and the section had a lot of blood and bubble flowing out,microscopic obsevation found bacillus with obtuse at both ends and spores,the bottom of the tube had loose and white precipitate in the broth,biochemical experiments showed that the isolate had acid-production and gas-production reactions which was unique to anaerobic bacteria causing Clostridium chauvoei infection,and 501 bp fragment was amplified by PCR.The results showed that two ill and dead cattle were casued by Clostridium chauvoei infection in Yanbian area,and proved the existence of Clostridium chauvoei.The test separated Clostridium chauvoei Yanbian strain for the frist time,which provided important reference basis for the prevention and treatment of Clostridium chauvoei infection,and laid the important foundation for the further study of drug and immune preventions of this disease.

This investigation aimed to examine coccidia infection in diary cattle in Yuzhong,Gansu province.An investigation was carried out on the Holstein calves less than one year old in six large-scale dairy farms.A total of 234 fecal samples were examined,and oocysts were identified to the species level on the basis of morphological features after positive samples were mixed thoroughly with 2.5% potassium dichromate solution.The results showed that the prevalence rate of coccidia in Holstein calves reached 46.15%.The average prevalence rate of the calves under 3 months old was 52.43%,the 4 to 6 months old calves was 37.50% and the 7 to 12 months old calves was 43.37%.Seven species of Eimeria coccidians and one species Isospora sp.including Eimeria bovis,Eimeria auburnensis,Eimeria zuernii,Eimeria alabamensis,Eimeria subspherica,Eimeria ellipsoidalli,Eimeria canadensisi,were identified in the investigation,and Eimeria bovis,Eimeria auburnensis,Eimeria zuernii were the dominant species.In conclusion,the prevalence rate of coccidiosis in dairy cattles in Yuzhong,Gansu province was high.Therefore,appropriate strategies and measures should be taken to control coccidiosis prevalence in dairy cows in this region.