Acta Veterinaria et Zootechnica Sinica

Origin, Evolvement and Resistance Mechanism of Polymorphism of MHC Molecules

CHEN Fang-fang;PAN Ling;GENG Zhao-yu;LIU Xue-lan;YU Wei-yi

2010, 41(9): 1061-1067. doi:

Abstract ( 774 )

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Cognition of MHC (Major histocompatibility complex)has been developed from only as an explanation of the phenomenon related with histocompatibility and exclusion to a gene group referred to resistance and all immune response at present. MHC gene is the ancestral sequences that existed before separation of species, and has evolved and generated significant diversity among species and among individuals in the same specie. The difference of gene structure appears among different species in a present or absent gene framework, or a highly plastic region or a single cola or multiple coli in genome or in an arrange manner and the amount of coli and alleles. A similarity of gene structure exists among relative species, however, difference has been found in the types of genomic rearrangement and pseudogene. A highly polymorphic alleles exhibit among individuals in the same specie and its genetic basic is point mutation based on some nucleotide substitution. The factors in environment, such as pathogen-mediated selection pressure are most likely to drive this process. In some research, the MHC haplotype or the locusspecific cluster with resistance to specific pathogen has been detected. The peptide binding region (PBR) in MHC is among the most polymorphic regions found in vertebrate taxa, while the affinity between PBR and antigen peptide results in disease resistance or susceptibility for animals. New techniques of molecular biology and bioinformatics provide available tools to investigate MHC polymorphism, especially to explore the mechanism of mutual action between MHC molecule and epitope.

Cloning and Expressional Analysis of PGC-1β and PRC cDNA in Pig

CHANG Zhi-guang;LU Rong-hua;JI Hong;SU Shang-shun;GAO Xiao-juan;YANG Gong-she

2010, 41(9): 1068-1075. doi:

Abstract ( 749 )

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The aim of this study was to clone the cDNA of PGC-1β and PRC gene from the heart of porcine by RT-PCR approach and to analyze its sequence and expression profile. According to the PGC-1β and PRC mRNA sequences of human which were the homologous sequences of pig searched in the EST library, the partial PGC-1β and PRC cDNA sequence of pigs were cloned by RT-PCR, and their expression profile were detected in heart, muscle, spleen, liver, lung, kidney, small intestine and adipose tissue by using RT-PCR and Western blot. The result showed that the PGC-1β and PRC were 1 251 and 1 254 bp in length, and encoding 415 and 417 amino acids(GenBank accession number：FJ377680 and FJ479491), respectively. Bioinformatic analysis found that the homology of nucleotide sequence and amino acid were about 80% compared with the counterpart sequences of other species, and containing TPPTTPP and DHDY conserved sequences, as well as the RNA recognition motif(RRM)conserved domains. PGC-1β was widely expressed in 8 tissues,while PRC was highly expressed only in heart, liver, skeletal muscle and kidney,but not detected in spleen and small intestine. The result suggested that the differention of expression level of the PGC-1β and PRC may be related to their physiological functions in different tissues.The results will be helpful for elucidating the physiological function and regulatory mechanisms of PGC-1 family.

Genetic Variation of the Lpin1 c.1727C>T and Its Association with Fattiness Traits in Chicken

WANG Bo-jun;SUN Ling;LI Shao-bo;ZHU Zhi-ming;LIU Bang

2010, 41(9): 1076-1081. doi:

Abstract ( 706 )

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Using genome scanning method, three missense mutation sites of Lpin1 gene located in the coding region were found, which were c.391G>A mutation(Asp131Asn), c.1727C>T mutation(Ala576Val) and c.2287A>C mutation(Met763Leu). Three pairs of primers were designed according to amplification-created restriction site PCR(ACRS-PCR)methods and were used to amplify the fragments covering the corresponding mutation. The PCR products were distinguished by the restriction fragment length polymorphism(RFLP)method. The association analysis between different genotypes and fattiness traits were performed in Tibetan chicken, Recessive White, crossbreeds of Tibetan♂×Recessive♀and Tibetan♂×(Tibetan♂×Recessive♀)populations. The genetic diversity about mutation sites was analyzed in 5 native chicken breeds(Gushi, Xiaoshan, Beijing Fatty, Silky Sooty and Chahua). The results showed that the polymorphism of the Lpin1 c.1727C>T mutation was apparently abundant in different chicken breeds while the others were monotonous. It revealed that the frequency of C allele in c.1727C>T mutation site was higher than that of A allele, and the distribution pattern of c.1727C>T locus genotype in the 5 chicken populations obeyed the Hardy-Weinberg equilibrium through the χ2 test, except the Gushi chicken and the Beijing Fatty chicken. The association analysis showed that there were a significant association between the c.1727C>T site polymorphism and subcutaneous fat thickness(P<0.05), and abdominal fat weight(P<0.01), and also revealed that the chickens with CC genotype had a higher subcutaneous fat thickness than chickens with the CT or TT genotype. Meanwhile, the chickens with TT genotype had a higher abdominal fat weight than chickens with other genotypes(CT or TT).

between mRNA Expression of Duck BLVRA Gene in Interstitial Part of Uterine Tube and Green Shell Character

ZHANG Bao-le;LI Guo-qin;ZHANG He;;SHEN Jun-da;LI Jin-jun;TAO Zheng-rong;CHEN Yi-chun;WANG De-qian;CHEN Wen-biao;SHI Fang-xiong;LU Li-zhi

2010, 41(9): 1082-1089. doi:

Abstract ( 1018 )

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In order to study the structure and function of biliverdin reductase A(BLVRA)gene and its correlation with the character of green shell, the unknown sequences of 3′and 5′ ends of duck BLVRA mRNA were cloned by using RTPCR and 5′RACE methods and four pairs of primers were designed and synthesized according to the known coding sequence(CDS)of duck BLVRA gene in the current project. Gene expression levels of BLVRA were measured by real-time quantitative PCR, and the eggshell colors were determined by the reflection coefficient method. The results showed that the cDNA sequence of BLVRA was comprised of 1 071 nucleotides, encoding 302 amino acids and having the isoelectric point of 7.15 and the molecular weight of 34.3 ku. The 5′ and 3′ ends of the duck BLVRA gene sequence published in the GenBank were extended 238 and 199 bp, respectively. Comparing the putative amino acid of duck BLVRA gene with that of other animals showed that it shared 95.3%, 95.0%, 70.0%, 61.1%, 60.5% and 59.5% homology with Gallus gallus, Taeniopygia guttata, Xenopus tropicalis, Bos taurus, Homo sapiens and Mus musculus, respectively, which proved that BLVRA gene was conservative in the process of evolution. The relative gene expression level of BLVRA in white eggshells of Jinyun duck was 3 times higher than that in dark green eggshells after normalization with GAPDH gene(P< 0.01). There was a significant positive correlationship between reflection coefficient of eggshell and BLVRA gene relative expression levels(r=0.719，P<0.05). Together, the result suggested that BLVRA gene could be used as a candidate gene for molecule breeding of the green shell duck.

Comparison of Plasma Protein Profiles from Perinatal Cow by Twodimensional Electrophoresis

NIU Li-li;LIU Tao;WEI Cai-hong;LI Hong-bin;DU Li-xin

2010, 41(9): 1090-1094. doi:

Abstract ( 1030 )

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In this study two-dimensional electrophoresis was applied to analyze the difference expression of plasma protein between preperinatal and postperinatal cows. The Hot-SDS method, pH 4-pH7 IPG strips(17 cm)and gradient gel were used for dairy cow plasma proteome analysis. Totally, 239 protein spots were detected using silver staining by PDQuest 6.0, which appeared repeatedly in three gels of each sample.12 spots showed more than 3 fold changes in abundance in plasma protein of cow. Plasma albumin and Haptoglobin were identified by MADILTOF/TOF.Their expressions were increased in the plasma of post-perinatal cow. Haptoglobin content ircreased in postperinatal cow plasma in this study, which showed the physiological changes of the postpartum cows.

Effect of α-MSH on Proliferation and Melanin Synthesis of Alpaca Skin Melanocytes in vitro

YU Zhi-hui;BAI Rui;FAN Rui-wen;YANG Gang;DONG Yan-jun;HE Jun-ping;DONG Chang-sheng

2010, 41(9): 1095-1101. doi:

Abstract ( 699 )

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To study the effects of α-Melanocyte Stimulating Hormone(α-MSH)on the proliferation and melanin synthesis of cultured Alpaca skin melanocytes．The cultured Alpaca skin melanocytes were treated with various concentration of α-MSH(0, 10^-9, 10^-8, 10^-7 mol·L^-1)in vitro. The effects of α-MSH on proliferation, melanin contents, MC1R, MITF and TYR gene expression of cultured melanocytes were observed. Three days after treatment with αMSH, the number of melanocytes increased and melanin contents, MC1R, TYR gene expression level also remarkably increased in treated cells compared to those in untreated cells(P＜0.05), especially when the concentration was 10^-8 mol·L^-1, and MITF gene also remarkably increased(P＜0.05)when the concentration was 10^-7 mol·L^-1.α-MSH can induce the dendrites elongated, proliferation, melanin synthesis and the expression of MC1R, MITF and TYR of alpaca skin melanocytes.

Production of MxA Transgenic Mice by Type A Spermatogonia-mediated Gene Transfer in vivo

JU Hui-ming;;BAI Li-jing;WANG Yi;PENG Da-xin;ZHANG Wei;HU Yan-xin;LIU Zong-ping;LI Kui

2010, 41(9): 1102-1108. doi:

Abstract ( 968 )

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The aims of this study were to explore the feasibility and stability of producing MxA transgenic mice mediated by Type-A spermatogonia in vivo. The pcDNA-MxA plasmid and transfection reagent ExGen500 were suspended and injected into testicle tissue of 7-day-old male ICR mice from 5 different points; the testis-mediated gene transfer(TGT)mice mated with wildtype female mice at different sexual maturity stages(6-, 12- and 24-week-old), to detect the integration and expression of foreign gene in offspring. The mice expressed MxA were chosen for challenge test, health status and respiratory system tissue pathological changes were observed. Detection result of MxA integration by PCR and Southern blot in the F₁-generation mice showed that foreign gene integration rates of 2 TGT mice at different sexual maturity stages(6-,12-, and 24-week-old)were 11.11%(2/18), 11.76%(2/17), 11.54%(3/26) and 13.64%(3/22), 13.33%(2/15), 11.76%(2/17),respectively. The average integration rates in two groups were 11.47% and 12.91%, the difference was not significant(P＞0.05). RT-PCR detection result showed that MxA gene was expressed in 3 of 14 integration mice. Infection challenge test with H5N1 influenza strains results showed that constitutional symptom and pathologic change degree of transgenic mice were more slighter than that of wild-type mice. The results indicated that the method of type-A spermatogonia mediated gene transfer shows bright future for application because it is a feasible and reproducible transgenic approach. MxA transgenic mice have disease resistance to H5N1 subtype avian influenza viruses to some degree.

Effects of Amino Acids on the Rumen Microbes Growth and Fermentation in vitro

WANG Hong-rong;XU Ai-qiu;WANG Meng-zhi;LI Shi-xia;WANG Huan-li;ZHANG Hong-wei

2010, 41(9): 1109-1116. doi:

Abstract ( 713 )

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This experiment was conducted to study the effects of AA on rumen microbes and fermentation in vitro. Four Xuhuai wether goats fitted with permanent ruminal cannulas were used to provide rumen liquor in vitro. The special amino acids removal method was applied. Treatments of total amino acid(Group A), Met-removal(Group B), Lys-removal(Group C), Branch-chain amino acid-removal(Group D), Aromatic amino acid-removal(Group E)were designed. The results of the study showed that the pH-value ranged between 6.01 and 6.45, and the highest mean value appeared in group C(6.23). Concentration of NH3-N ranged from 7.98 to 29.05 mg·100 mL^-1, with the highest mean value occuered in group A(18.16 mg·100 mL^-1). The order of the restricted role of AA-specific default on rumen micro-organisms(bacteria and protozoa)was that: BCAA> ARAA>Methionine> Lysine. The highest of P/B ratio was group B(101.37%), and the lowest was group D(88.22%). The highest of microbial AA-N ratio, bacteria AA-N, protozoa AA-N was group E, group D and group E, respectively, and the lowest of them was group C. Microbial diversity was demonstrated in SSCP fingerprint clearly, which revealed that the structure of bacteria or protozoa community was altered by substrates. Amino acid influences both on fermentation and microbial community in vitro.

In Vitro Fermentation Kinetic Analysis of the Carbohydrate Fractions in Various Types of Corn Stalks

XUE Hong-feng;REN Li-ping;MENG Qing-xiang

2010, 41(9): 1117-1125. doi:

Abstract ( 944 )

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The objective of this experiment was to evaluate the fermentation kinetics of The Cornell Net Carbohydrate and Protein System(CNCPS)carbohydrate fractions(A, B1, B2 and NDS)of various corn stalks′ types, including two varieties of high oil corn(HOC 647 and HOC 298), two varieties of typical corn(CAU 80 and CAU 3138)and one specific fodder corn(FC). The results showed that carbohydrate fractions were significantly different(P<0.001)among various corn stalks′ types. Contents of carbohydrate fractions A,B1 and NDS of HOC were highest among three types′ corn stalks, and gas productions of the three carbohydrate fractions from various corn stalks′ followed a similar order of their fraction contents (P<0.000 1). Contents of carbohydrate fraction B2 of HOC were lowest among three types′ corn stalks. These results indicated that HOC had better the nutritional value, because content of soluble carbohydrate of HOC was higher than that of other two corn stalks.

The Comparative Analysis of the Whole-genome Sequences of Mycobacterium tuberculosis and Mycobacterium bovis

ZHU Ru-yi;PAN Wen;ZHAO Ming-qiu;JU Chun-mei;YI Lin;WANG Jia-ying;HU Yong-ming;CHEN Jin-ding

2010, 41(9): 1126-1132. doi:

Abstract ( 726 )

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The aim of present study was to reveal the comparative differences of whole-genome sequences between Mycobacterium tuberculosis and Mycobacterium bovis, and to provide new molecular markers for identification and diagnosis of these two bacilli. The whole-genome sequences of M. tuberculosis H37Rv and M. bovis AF2122/97 were sequenced and their genomic comparison was performed by on-line tools. M. bovis AF2122/97 shared 99.95% genomic nucleotides identity with M. tuberculosis H37Rv. 14 large fragment deletions varied in a range from 0.8 to 12.7 kb and 6 small deletions varied from 12 to 714 bp were detected in AF2122/97 strain by comparison with H37Rv, while 6 large deletions ranged from 1.3 to 5.4 kb and 5 small deletions ranged from 10 to 48 bp, in turn, were discovered in H37Rv strain. The large fragment deletions of RD18, RD19 and RD20 in AF2122/97 strain, and all small deletions both in AF2122/97 and H37Rv strains were firstly reported. Deletion has led to a reduced genome size of M. bovis. Most of the deleted genes are found to be responsible for encoding cell wall components and secreted proteins, indicating that deletions of these genes may contribute to hostbacillus interaction or immune evasion. The new understanding might be helpful in the development of new drugs, diagnostic reagents and vaccines.

Infection of Low Pathogenic Avian Influenza in Poultry in Eastern China

ZHAO Guo;ZHAO Kun-kun;LU Xin-lun;XU Quan-gang;PENG Da-xin;LIU Xiu-fan

2010, 41(9): 1133-1137. doi:

Abstract ( 970 )

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From October 2008 to September 2009, 2 420 cloaca swab samples were collected from domestic poultry in the LPMs (livepoultry markets), and 83 strains of AIVs were isolated with an isolation rate of 7.36％. These strains of AIV belonged to 7 HA subtypes, from high to low, the order of the isolation rate was H6, H3, H4, H9, H11, H1, H8. The isolation rate of AIVs was distributed with significant seasonal variations, higher in winter and spring and lower in summer. The distribution of HA subtypes of AIVs was also uneven in different month. Waterfowl are considered as the important host of influenza A viruses, especially domestic ducks. So far, 8 HA subtypes (H1, H3, H4, H6, H8, H9, H10 and H11) and 7 NA subtypes (N1, N2, N3, N4, N5, N6 and N8) were isolated from domestic ducks. Strains of HA and NA subtypes assembled 18 subtypes. Domestic duck is still the most prone to mixed infections, and H6 subtypes played the main role, H3 and H9 were the other main mixed infection subtypes, providing a good carrier to generate a new subtype and virulence variation.

Genetic Variations of Main Structural Genes of Infectious Bronchitis Virus Strains Isolated in Guangxi Province of China

MO Mei-lan;LI Meng;WEI Ping;FAN Wen-sheng;HUANG Bai-cheng;LANG Ya-hui;CHEN Qiu-ying;HOU Jin-lian;WEI Tian-chao

2010, 41(9): 1138-1146. doi:

Abstract ( 635 )

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In order to study characteristics of genetic variations of avian infectious bronchitis virus (IBV) in Guangxi Province of China, spike S1 protein, nucleocapsid （N） and membrane （M） protein genes of 7 IBV strains isolated in Guangxi between 2004 and 2007 were amplified by reverse transcriptase polymerase chain reaction (RT-PCR), cloned, sequenced, compared and phylogenetically analyzed. The results showed that many point mutations occurred within the S1 gene in all viruses and gene insertions and deletions occurred in majority of isolates. Amino acid sequence homology of S1 gene among the 7 IBV isolates varied from 74.2% to 98.7%. Deletion and insertion were not found in the N genes, but point mutations were found. Amino acid sequence homology of N gene among the 7 IBV isolates varied from 91.7% to 99.3%. Mutations and insertions occurred in M genes. Amino acid sequence homology of M gene among the 7 IBV isolates varied from 90.7% to 98.2%. Majority of Guangxi IBV isolates showed variations in S1, N and M genes comparing with H120 vaccine strain and S1 gene showed the biggest variation among the isolates. 7 IBV isolates were clustered into 2, 2 and 3 groups in the phylogenetic trees based on the deduced amino acid sequences of S1, N and M genes. The phylogenetic trees based on the N and M gene sequences do not follow closely the phylogenetic clustering based on the S1 gene for 4 out of 7 isolates. These results suggested that the variations were occurred in majority of Guangxi IBV isolates at S1, N and M genes. Point mutations, insertions and deletions had contributed to the genetic diversity and emergency of IBV variants in Guangxi Province. The present results indicated that the genetic variations may provide an explanation for the reasons of vaccines can’t protect effectively against epidemic strains in the field.

Construction of Recombinant Baculovirus Containing Peste des Petitis Ruminants Virus N Gene and Establishment of Indirect ELISA for Detecting the Serum Antibody

LI Wei;LI Gang;WU Xiao-dong;QIU Wen-ying;ZHANG Kun;Hermann Unger

2010, 41(9): 1147-1153. doi:

Abstract ( 1135 )

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This experiment was conducted to diagnose Peste des Petitis Ruminants based on the eukaryo-expressed PPRV nucleoprotein through indirect ELISA. The full-length PPRV nucleoprotein gene was obtained from viral genome RNA by RT-PCR. The amplified fragments were cloned into baculovirus donor vectors pFastHTA of Bac-to-Bac system. These recombinant plasmids, pFastHTA-PPRV-N, were transformed into DH10Bac host bacteria to get recombinant shuttle plasmids, pBacmid-PPRV-N. Recombinant baculovirus, Bacmid-PPRV-N, was generated for expressing PPRV nucleoprotein by transfecting recombinant pBacmid-PPRV-N with LipofectAMINE 2000 into Sf21 insect cells. PPRV nucleoprotein which efficiently expressed by baculovirus in Sf21 cells was verified by SDS-PAGE and Western blot. Furthermore, indirect ELISA was developed using recombinant PPRV nucleoprotein as coating antigen. 37 goats′ sera from epidemic area in Tibet Autonomous Region and 92 goats′ sera from non-infected area in Qinghai Province were detected by indirect ELISA and international standard cELISA kit simultaneously. The sensitivity and specificity of indirect ELISA were 100% and 96.2% compared with cELISA kit. The coincidence rate of the two methods was 96.9%. The results demonstrated that the indirect ELISA established in this study works well in the PPR diagnosis.

Identification and Molecular Characteristic Analysis of Orf Virus (ORFV) Hubei Strain

ZHANG Ke-shan;HE Ji-jun;SHANG You-jun;GUO Jian-hong;ZHENG Hai-xue;TIAN Hong;JIN Ye;LIU Yong-jie;WANG Guang-xiang;LIU Xiang-tao

2010, 41(9): 1154-1157. doi:

Abstract ( 1006 )

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To identify suspected goats Orf case and analyze the molecular characteristics of Orf Virus (ORFV), crust materials of lambs with clinical sore mouth symptom from a farm in HuBei Province were used and researched. ORFV in tissue scrapings from the lips was confirmed by electron microscope, HE stain and specific gene(B2L)PCR amplification. Molecular characterization of ORFV strain was performed using amino acid sequence homology comparison and phylogenetic analysis based on B2L gene. The results showed that this disease was caused by ORFV(named TS/HuB/CHIA/2009), B2L gene sequence was submitted to GenBank and obtained accession number GU320351. Phylogenetic tree analysis demonstrated that the GU320351 has a closest genetic relationship with DQ904351, which was isolated from Taiwan Province of China in 2007. And GU320351 was in the evolutionary branch contain DQ263306, DQ263305, DQ263304 and DQ263303, which was isolated from India in 2004 and 2005. Research of this paper accumulated useful material for further studying of molecular epidemiology of ORFV in China.

Effects of Different Expression Vectors and Temperatures on the Activityof Eimeria tenella Malonyl-CoA:ACP Transacylase

SUN Ming-fei;QIN Zong-hua;XIE Ming-quan;YUAN Jian-feng;LV Min-na;YU Jin-shu;WU Cai-yan;CAI Jian-ping

2010, 41(9): 1158-1165. doi:

Abstract ( 722 )

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This experiment was conducted to study the effects of different expression vectors and temperatures on the activity of recombinant Eimeria tenella Malonyl-CoA:ACP Transacylase (rEtMCAT). Using the ORF sequence encoding EtMCAT predicted by bioinformatic analysis as a template, 3 pairs of primers were designed in order to amplify the following fragments: the entire ORF, the ORF without N-terminal signal peptide, the ORF without N-terminal signal peptide and transit peptide，respectively. Each of the 3 cloned fragments was individually inserted into 3 plasmid vectors pMAL-c2x, pET-32a(+) and pProExHTa, respectively. All the recombinant vectors were transfected into E. coli Rosseta(DE3), and induced with IPTG under different temperature conditions (37, 30 and 16 ℃), respectively. The enzymatic activities of rEtMCAT were measured using the a-ketoglutarate dehydrogenase (KDH)coupled assay system. The results revealed that only the ORF encoding the mature peptide, that is, excised the N-terminal signal peptide and transit peptide could be expressed in 3 vector systems, and high content of soluble rEtMCAT was only expressed in recombinant plasmid pMAL-c2x-mcat or pET32a(+)-mcat, in which their expressed products had a similar enzymatic activity. The lower culture temperature (at 30 and 16 ℃) can enhance their expression. But there was no any soluble protein expressed in Rosseta(DE3)/pProExHTa-mcat, and the recombinant protein expressed in inclusion body was no enzymatic activity even after renaturation. The characterizations of expression vectors used in experiments, especially the type of tag protein, as well as the expression temperatures have significant effects on the production and enzymatic activity of rEtMCAT expressed in E. coli Rosseta(DE3). It is suggest that the rEtMCAT expressed in inclusion body can not be used for the enzymatic assay.

Cloning， Expression of Bovine Tracheal Defensin in E.coli and Its Antibacterial Activity and Tissue Distribution

GAO Xiao-yan;WANG Chang-fa;LIU Shun-de;LI Qiu-ling;YANG Hong-jun;YANG Shao-hua;ZHONG Ji-feng;HE Hong-bin

2010, 41(9): 1166-1171. doi:

Abstract ( 695 )

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The study was conducted to clone bovine tracheal antimicrobial peptide gene (TAP) from bovine tracheas, and recombinant bovine TAP protein was expressed in E. coli. Its antimicrobial activity were determined. In addition, the tissue distribution of the TAP gene in bovine was investigated. The mRNA of bovine TAP was cloned from tracheas of Holstein cow by RT-PCR. Based on the analysis of the sequence of bTAP and the codon preference in E.coli expression system, the bovine TAP gene was synthesized and inserted into vector pET32a(+) to construct plasmid pET32a(+)-TAP. The plasmid was expressed in E. coli BL21(DE3) cell by induction of IPTG. The recombinant fusion protein was purified, and determined by SDS-PAGE and Western Blot. Antimicrobial activity of the recombinant fusion protein were measured by filter paper method. Tissues distribution of the TAP gene has been demonstrated in bovine by RT-PCR. The full length cDNA of bovine TAP was consisted of 114 bp, encoding 38 amino acid residues. Western Blot results showed that the recombinant proteins were expressed in E. coli with molecular weight of 24 kD, exists in soluble form and accounted for 52.1% of the whole bactera proteins. A single band appeared in Western Blot detection. The content of 0.115 μg·μL-1 of the purified TAP protein had antibacterial activity against bovine Sta.Aureus in vitro. Bovine TAP mRNA was widely expressed in the tissues of bovine, such as throat, lungs, trachea, small intestine, liver, heart, oral mucosa, and it was micro-expressed in the nasal mucosa, lingual mucosa and almost not expressed in the skin. The results showed that bovine TAP protein was highly expressed in E. coli, and can be used for transgenic dairy cow which anti-mastitis and developing the product of genetic engineering in future.

Effect of Monochromatic Light on Expression of c-Fos in the SCN and SGCof Optic Tectum in Broiler

JIN Er-hui;JIA Fei;YANG Guang;WANG Zi-xu;CHEN Yao-xing

2010, 41(9): 1172-1176. doi:

Abstract ( 631 )

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This study was conducted to compare the effect of monochromatic light on expression of c-Fos in the SGC of optic tectum and SCN in broilers. Forty onedayold male AA (Arbor Acres) broilers were respectively reared under four light treatments, white (400-760 nm), red (660 nm), green (560 nm), blue (480 nm) by using LED (light-emitting diodes) as light sources until the 14th day.Light intensity was 15 lx. Light exposure was scheduled for 20 hours of light and 4 hours of darkness during the entire experiment. The results showed: in the SCN, c-Fos expression of BL group were significantly lower by 29.76%， 17.88% and 22.23% than that of WL, RL and GL groups respectively (P<0.05), no significant difference were found among WL, RL and GL groups (P>0.05). Similarly, c-Fos expression of BL group were significantly lower by 17.78%， 21.75% and 23.63%than that of WL, RL and GL groups, respectively (P<0.05), no significant difference were also found among WL, RL and GL groups in the SGC of optic tectum (P>0.05). These results suggested that blue light suppress c-Fos expression of neuron in the SCN and SGC of optic tectum compare to red and green light to some extent.

Localization, Distribution and Development of Ghrelin-immunopositive Neurons and Influence of Fasting on Them in Brain of Broiler Chickens

YE Yuan-lan;LI Yu-gu;WEI Feng-mei;MA Yong-jiang;ZHANG Yuan;JIANG Qing-yan

2010, 41(9): 1177-1184. doi:

Abstract ( 692 )

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In this paper, the localization and distribution of ghrelin-immunopositive neurons in the brain, the development of these neurons in the major nucleus of hypothalamus in 2, 16, 30, 44 and 58-day-old, and the influence of fasting 12, 24, 48 h and refeeding 4 h in 30-day-old on these neurons in broiler chickens were investigated by immunohistochemical staining and microscopic image analysis. The results showed that the ghrelin-immunopositive neurons were observed in the nucleus periventvicularis accuatas, the nucleus paramedianus, the nucleus medialis, the area periventricularis, the area rostromedialis and the area rotrolateralis hypothalami, the nucleus rotundus and the nucleus ovoidali, the gray matter of colliculus mesencephali and the nucleus ruber, the nucleus vestibularis ventrolateralis, the nucleus cerebellaris medialis and the granular layer of cerebellar cortex, and the polymorphic layer of cerebral cortex; with the chicken growth, the number of these neurons was increased, their immunopositive reaction level was enhanced, but their density was decreased; during the fasting and refeeding, their immunopositive reaction level was weakened and their density was decreased.

Effects of Water Soluble Alfalfa Polysaccharides on Cell Proliferation of Broiler Lymphocytes in Vitro

LIU Qing-xue;DONG Xiao-fang;TONG Jian-ming;XU Chun-yan;WANG Jun-li;ZHANG Qi

2010, 41(9): 1185-1190. doi:

Abstract ( 652 )

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To investigate the effects of Water Soluble Alfalfa Polysaccharides(WSAP)on cell proliferation of broiler lymphocytes in vitro. Different concentrations of WSAP were added into cultured with ConA stimulated chicken peripheral blood, thymus, spleen lymphocytes and into cultured with LPS stimulated chicken peripheral blood, spleen, bursa of fabricius, cultured for 48 hours, use MTT colorimetric determination of lymphocyte proliferation. The results showed that(1)In this experiment, the OD values of T, B lymphocytes in a certain concentration of WSAP (4-20 μg·mL^-1)were higher than those in control groups, with the polysaccharide concentration increased, OD values increased first(1-20 μg·mL^-1)and then decreased(100-200 μg·mL^-1);(2)The same concentration WSAP on broiler peripheral blood and other immune organs, the stimulus intensity were different, peripheral blood and spleen T, B lymphocytes OD values were less than the thymus and bursa lymphocytes OD values;(3)The best stimulus concentration for peripheral blood and other organs were difference, for peripheral blood was 20 μg·mL^-1, for spleen was 8 μg·mL^-1， for bursa and thymus with 15 μg·mL^-1. In this experiment, the concentration of WSAP with 8, 15, 20 μg·mL^-1 could promote the broiler lymphocyte proliferation, which may be the one of the mechanisms that polysaccharides could enhance immune function.

Differential Proteomics Analysis of Plasma Protein from Escherichia Coli Infected and Clinical Healthy Dairy Cows

YANG Yong-xin;CHENG Guang-long;ZHAO Hui-ling;JIANG Xi-chun;CHEN Sheng

2010, 41(9): 1191-1197. doi:

Abstract ( 1045 )

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Escherichia coli are most common pathogen from a contaminated environment in lactating dairy cows. For investigating the changes of plasma protein form E. coli infected cows and exploring the mechanism of host defense, such changes were examined using an integrated proteomic approaches consisting of plasma protein and minor abundance protein enrichment by ProteoMiner kit, protein separation by two-dimensional gel electrophoresis, stained with Coomassie Blue G-250 solution, and protein identification by HPLC equipped with ion trap mass spectrometer. The results showed that the nineteen protein spots were changed in cows infected with E. coli mastitis, and fifteen spots were identified as seven proteins. Haptoglobin, alpha 1 acid glycoprotein, and serum amyloid protein A were up-regulated in plasma from E. coli infected cows. Expression of haptoglobin by ELISA method was significantly higher in E. coli infected cows than in healthy controls(P<0.01), E. coli infected cows altered the plasma protein level, and suggested that changes of proteins may be useful to clarify the host defense mechanisms and response to invasion of mammary tissues by E. coli infection, and potential protein targets for diagnose and treatment.

Immunosuppressive Characters of Avian Leukosis Virus Subgroup J

ZHANG Li;LIU Qing;WANG Xiao-wei;WANG Feng;CHEN Hong-bo;CHENG Zi-qiang

2010, 41(9): 1198-1202. doi:

Abstract ( 700 )

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SPF birds were inoculated with avian leukosis virus subgroup J- NX0101 strain at 1 day age and 7 day age, respectively-analog congenital infection and early infections. Detect the effect of immune system caused by ALV-J. Body weight, immune organs weight, histopathology, blood cells, CD4⁺ and CD8⁺T lymphocytes in spleen and thymus were tested. Results showed that ALV-J suppressed significantly not only body weight but also immune organs, especially in thymus and the bursa of Fabricious from 2 weeks age. The most serious immunosuppression occurred post inoculation 3 to 4 weeks. Histopathology observation found that most immune cells were destroyed and interstitial connective tissue proliferation in thymus and the bursa of Fabrisious. The other organs showed inflammatory infiltrate and hemorrhage. Tumor was not seen in any organs. Blood detection showed that ALV-J caused granular leukocytes, lymphocytes and erythrocytes decreasing, especially group 1. CD4⁺ and CD8⁺T lymphocytes detection in spleen and thymus showed that CD4⁺ cell number decreased, while CD8⁺ cell number increased at 5 weeks age. It is confirmed that the immunosuppression caused by ALV-J was high related with the two kinds of cells. The study demonstrated that the latent period of ALV-J in vivo was at 3 to 4 weeks after infection.

Study on Intrauterine Environment during Early Pregnancy of Hu Sheep

MAO Da-gan;SHI Guo-qing;;ZHANG Hong-lin;YAN Yu-qin;CHENG Rui-he;YANG Li-guo

2010, 41(9): 1203-1207. doi:

Abstract ( 672 )

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To study the fecundity mechanism of Hu sheep，the intrauterine environment during early pregnancy (Day 43) of Hu sheep (Hu) were examined compared to Xinjiang Fine Wool sheep (XJ)． The fetal average weight of Hu was lower than that of XJ (P<0.05) and there was no difference in fetal length and fetal membrane weight (P>0.05)．The uterine weight and the outer and inside diameter of uterine gland of Hu were significantly higher than that of XJ (P<0.01)，while the thickness of mesometrium of Hu was significantly lower than that of XJ (P<0.01)．The RNA/DNA ratio and dry material content in cotyledon of Hu were significantly higher than that of XJ (P<0.01)，while the glucose content was significantly lower than that of XJ (P<0.01)，and the number and size of cotyledon had no significant differences (P>0.05) between the two breeds． The results suggested that the fecundity of Hu sheep was possibly related to the smaller embryo in the larger uterine antrum，the abundant nutrients in the larger uterine gland cavity，the fine transportation function of active cotyledon cells and so as to protect embryos to develop all right．

Analysis of Genetic Diversity in Ritu Tibetan Goats by ISSR

WANG Yong;WANG Jie;XU Qi-shu;ZI Xiang-dong;OUYANG Xi;LIU Lu-shu;XIAO Yi-xi

2010, 41(9): 1208-1212. doi:

Abstract ( 1049 )

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Inter simple sequence repeat(ISSR)was used to analyze the genetic diversity of Tibetan goats population in Ritu county. Ten primers were selected from 93 ISSR primers and then used to detect the genetic diversity in 107 Tibetan goats. The obtained data were analyzed by using Excel 2003 and SPSS11.0 softwares. Ten out of 93 ISSR primers produced 112 DNA bands, among them 75 bands were polymorphic markers and the size of amplified fragment ranged from 219 to 2 534 bp. It indicated that the selected primers were specific and could produce enough polymorphic bands. The proportions of polymorphic loci(P)in Ritu Tibetan goats was 66.96%. The genetic similarity index(s)within the detected population ranged from 0.460 0 to 0.740 5(with average of 0.564 7). The averaged individual genetic difference(D)was 0.435 3. The gene frequency(f)ranged from 0.045 5 to 1.000 0. These results of ISSR analysis indicated that the Ritu Tibetan goats population presents high level of genetic diversity and genetic variations existed among individuals.

Observation of the Histological Structure of the Yak（Bos grunniens） Uterus during Estrus Cycle

DUAN Yong-xia;CUI Yan;YU Si-jiu;YANG Bo

2010, 41(9): 1213-1218. doi:

Abstract ( 695 )

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Histological and morphometric methods were used to study the changes of histology and morphology of yak′s uterus during estrus cycle. The results showed that the wall of yak′s uterus was composed of mucous membrane(endometrium), tunica muscularis(myometrium)and tunica adventitia(perimetrium). The endometrial surface and glandular epithelium were simple columnar epithelium at follicular phase and increased obviously at luteal phase which were pseudostratified columnar epithelium. The endometrial surface epithelium consisted of secretory cells and a few lymphocytes. The secretory cells included light and dark cells. In general, the cytoplasm of the light cells was lighter than that of dark cells, and has a large and columnar shape. The dark one has a laterally compressed and slim shape. The component of the glandular epithelium was similar to that of the surface epithelium. There were abundance of PAS-positive granules in the endometrial surface epithelium, glandular epithelium and stroma were observed with PAS reaction during the luteal phase more than that of the follicular phase. The invaginated(telescoping)gland, enlarged gland and accumulated lymphocytes were occasionally found in the lamina propria during the follicular phase, but during the luteal phase only the telescoping gland was observed in the lamina propria. Furthermore, the thickness of endometrium and myometrium increased significantly at luteal phase, than that of the follicular phase(P＜0.01). The mean gland duct diameter, area, perimeter, total glandular area and perimeter of the endometrial glands were significantly higher at the luteal phases than that at the follicular phase(P＜0.01), but there was no significant change in the number of gland ducts(P>0.05). Therefore, there was a regular change in the histological structure of the yak′s uterus during the estrus cycle.

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