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24 August 2012, Volume 43 Issue 8
Research for Marginal Profits of Growth and Reproduction Traits of Pigs in Beijing Area
SHI Xiaoming;DING Xiangdong;ZHANG Qin;XIAO Wei
2012, 43(8):  1177-1185.  doi:
Abstract ( 322 )   PDF (605KB) ( 907 )  
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This study was conducted to estimate the marginal profits of growth and reproduction traits of pigs in Beijing area. A farm model was constructed using balancing method and system analysis. The production of commercial pigs was simulated, and the related data was obtained from farms. The results showed that the marginal profits of production and reproduction traits were 20.5 RMB for litter size born alive(pig/litter), -0.3 RMB for days at body weight 100 kg(day), 0.04 RMB for average daily gain(g·d-1), -186.4 RMB for feed conversion efficiency(kg·kg-1),-15 RMB for carcass backfat thickness(mm), and 1.8 RMB for sow reproductive longevity(litter). These results indicate that changes of the market prices affect the marginal profits of the traits investigated greatly.
遗传繁育
Distribution Patterns of GnRH and GnRHR Positive Cells in Hypothalamus at Different Postnatal Stages in Jining Gray Goats
LIU Xiao;WANG Shuying;YIN Yutao;ZHANG Demin; HUANG Libo;HOU Yanmeng;
2012, 43(8):  1186-1191.  doi:
Abstract ( 312 )   PDF (2060KB) ( 418 )  
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In order to detect the morphological changes and distribution patterns of GnRH and GnRHR in hypothalamus at different postnatal stages in Jining Gray goats, immunohistochemical Strept Avidin BiotinPeroxidase Complex (SABC) threestep method was used to study the distribution of GnRH and GnRHR in hypothalamus in goats at 0, 2, 4 and 6 months old. The results showed that GnRH and GnRHR positive cells were widely scattered in hypothalamus. Most of the positive cells were distributed in medial preoptic area (MPA), mamillary nucleus (MB), supraoptic nucleus (SON) and suprachiasmatic nucleus (SCN). The size and number of positive cells were increased with the increase of month old. The numbers of positive GnRH and GnRHR cells were significantly higher at 2 month old (puberty) than that at birth. The results suggest that GnRH and GnRHR secreted by hypothalamus might play a pivotal role in the onset of puberty and the maintenance of maturation in Jining Gray goat at the postnatal developing stage.
Construction and Detection of the Adenoviral Vector Encoding Zincfinger Nucleases Targeted to MSTN in Sheep
ZHANG Cunfang;WANG Ling;REN Gang;ZHANG Tingting;WANG Rui; YAN Guoyong;ZHANG Zhiying
2012, 43(8):  1192-1199.  doi:
Abstract ( 342 )   PDF (1260KB) ( 556 )  
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The aim of the study was to knock out the MSTN gene in sheep, and adenoviral vector encoding zincfinger nucleases (ZFN) targeted to the exon 1 of MSTN was constructed and packaged. Based on the high transfection efficiency and nonintegration of the adenovirus combinding with the specificity and high performance of the ZFN, disruption of the MSTN in sheep was expected in the further research. DNA sequences of T2A and the two designed ZFNs monomers were amplified by PCR and inserted into the pAdTrackCMV successively. The positive ones got were named as pAdTrackZFNLT2AZFNR. Then pAdTrackZFNLT2AZFNR and pAdEasy1 were cotransfected into E. coli. BJ5183 in which homologous recombination would happen between the two plasmids, the recombinant adenoviral vector was named as pAdEasyZFNLT2AZFNR. The recombinant vector was transfected into the HEK293 subsequently to produce the final adenovirus. Finally, the adenovirus was identified by PCR and the titer was detected, then the positive ones were reduplicated. The final adenovirus were added to the culture medium of sheep fetus fibroblasts to further confirm the infectivity of the virus and identify the activity of ZFN in mammalian cells by Western blot. The adenoviral vector containing MSTNZFN expression cassette was verified by enzymatic digestion and sequencing, and ZFN could recognize and cleave the target site of MSTN. The adenoviral vector encoding ZFN could target to the MSTN in sheep, and it was constructed successfully.
Association of IRS-1 Gene Polymorphism with Body Measurement and Meat Quality Traits in Qinchuan Cattle
MA Xianghui;ZAN Linsen;;GAO Jianbin;YANG Ning;ZHU Guangxing;CHENG Gong;WANG Hongbao;HAO Ruijie;FU Changzhen;JIANG Bijie; ZHAN Xiaoli;BAI Yinping
2012, 43(8):  1200-1209.  doi:
Abstract ( 376 )   PDF (934KB) ( 530 )  
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The aim of this study was to study the polymorphisms on exons of IRS1 gene and their association with several body measurements and meat quality traits in Qinchuan cattle. The polymorphisms on exons of bovine IRS1 gene was detected by using PCRSSCP and DNA sequencing technology. 384 Qinchuan cattle with 18 to 24 months old were selected and the polymorphisms of bovine IRS1 gene and their associations with body measurements and meat quality traits were analyzed. Two SNPs on exons of bovine IRS1 gene were identified: B1 site g. +2346 G>A, Glu>Glu; B2 site: g. +2394G>A, Leu>Leu. Two genotypes at each sites( B1 site: CC and CD;B2 site:AA and AB) were identified and homozygote DD and BB were not detected in the study. The result of χ2 test showed that B1 site was in agreement with the HardyWeinberg equilibrium (P>0.05), B2 site was significantly deviated from HardyWeinberg equilibrium (P<0.01). At B1 site, genotype CC was dominant genotype, C was preponderant allele, the site belonged to intermediate genetic diversity. At B2 site, genotype AA was dominant genotype, A was preponderant allele, the site belonged to low genetic diversity. The result of association analysis demonstrated, at B1 site, individuals with different genotypes had significant differences in body length, hip width, chest circumference and loineye area (P<0.05), at B2 site, individuals with different genotypes had significant differences in hip width, chest depth and backfat thickness (P<0.05). Furthermore, haplotype analysis showed that body measurement traits of individuals with combined genotype ABCD were significantly higher than those of the other kinds of combinations except on backfat thickness and intramuscular fatty content. The effect of multiloci combinations on individuals were higher than those of singlemarker loci. As the result showed, IRS1 gene had a significant effect on body measurement and carcass traits in Qinchuan cattle. Hence, it could be took as a candidate gene for selecting new breed of Qinchuan cattle and the ABCD genotype maybe the best one for the Qinchuan cattle breeding.
Purification and Immunogenicity of Matured Peptide of Recombinant Myostatin in Yak
MAO Liang;XU Yaou;ZHENG Yucai
2012, 43(8):  1210-1214.  doi:
Abstract ( 311 )   PDF (478KB) ( 520 )  
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In order to understand the action mechanism of yak MSTN, seek for possible method to block its activity and finally realize an efficient growth in yak, MSTN gene was recombinantly expressed in prokaryotic cell and purified for testing its immunogenicity. Firstly, yak MSTN mature peptide gene was cloned using specific primers and reversetranscription PCR method. Secondly, the target fragment was cloned into pET28a(+) vector and then transformed into E. coli BL21 (DE3) for prokaryotic expression. The target peptide was purified from total prokaryotic expression products. Finally, immunogenicity of yak MSTN peptide was detected by ELISA. The MSTN mature peptide gene inserted into the recombinant expression vector contained 330 bp and encoded 109 amino acids in yak. The expression host transformants were induced by adding 0.1 mmol·L-1 IPTG to the LB medium, and the content of target protein was about 21% of the total proteins after six hours of induction. The highly purified yak MSTN mature peptide was obtained by affinity chromatography and its molecular weight was 16.5 ku. The result of ELISA showed that the mature yak MSTN recombinant peptide had an excellent immunogenicity and the titer of rabbit serum antiyak MSTN was 1∶32 000. The study of highly efficient expressed and purified yak mature MSTN peptide with excellent immunogenicity obtained from this study paved a way to study its functional mechanism that may be further explored to improve the meat production performance of yak.
Effects of Free Radical Absorptive Antioxidants on the Development of Somatic Cell Nuclear Transfer Yanbian Yellow Cattle Embryos
YIN Duo;ZHU Yuxia;LIU Haixing;LI Zhongshu;FANG Nanzhu
2012, 43(8):  1215-1221.  doi:
Abstract ( 260 )   PDF (398KB) ( 439 )  
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In order to improve the development efficiency of somatic cell nuclear transfer embryos of Yanbian Yellow cattle, free radical absorptive antioxidants, such as vitamin E, EGCG and sodium selenite, were added in culture medium in vitro with different concentrations for observing the development of the reconstructed embroys. Oocytes were cultured 2022 h in maturation medium, oocytes with first polar body were chose for enucleation, injection, integration and activation, then restructured embryos were cultured in in vitro culture mediums with antioxidants with different concentrations. On the cleavage rate, in VE study, results showed that 100 μmol·L-1 group was significantly higher than those of other concertrations(84.58% vs 57.64%,70.87%,67.64%);there were no significant differences between 50 and 200 μmol·L-1 groups,but they were both significantly higher than that of 0 μmol·L-1 group(70.87%,67.64% vs 57.64%);In EGCG study,results showed that 7.5 and 15 μmol·L-1 groups were significantly higher than that of 0 μmol·L-1 group(79.47%,81.67%vs 69.47%)and there were no significant differences between other groups;In SS study,5 and 0 ng·mL-1 group were significantly higher than 10 ng·mL-1 group(80.44% vs 70.27%,67.16%)while there were no significant differences with 2.5 ng·mL-1 group(80.44% vs 73.92%), and there were no significant differences between other groups.On the Blastocyst rate,in VE study,results showed that 100 μmol·L-1 group was significantly higher than those of other concertrations(26.36% vs 13.04%,18.08%,17.26%);In EGCG study,results showed that 15 μmol·L-1 group was significantly higher than 0 and 30 μmol·L-1 groups (24.86% vs 8.99%,12.10%)and there was no significant differences with 7.5 μmol·L-1 group;In SS study, 5 ng·mL-1 group was significantly higher than that of 0 and 10 ng·mL-1 groups(23.31% vs 10.32%,12.65%);2.5 ng·mL-1 group was significantly higher than that of 0 and 10 ng·mL-1 groups(19.97% vs 10.32%,12.65%) while there were no significant differences between other two groups. Adding 100 μmol·L-1 vitamin E,15 μmol·L-1 EGCG or 5 ng·mL-1 sodium selenite alone in vitro into culture medium can enhance the early development rate of Yanbian Yellow cattle restructured embryos in vitro.
Effects of Ecadherin Antibody on Mouse Early Embryo Development
WU Xinglong;WANG Mianjuan;HU Chunchao;WANG Pengbo;LI Xiangyun
2012, 43(8):  1222-1229.  doi:
Abstract ( 282 )   PDF (5074KB) ( 563 )  
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To investigate the effects of Ecadherin antibody (ECCD1) on mouse embryo development, ECCD1 antibody with different concentrations was added into KSOM media when mouse 8cell embryos were cultured in vitro, and then the developmental potential of ECCD1treated embryos were detected by morphology, alkaline phosphatase activity, pluripotent marker (SSEA1, Oct4, Sox2, Nanog) immunostaining, attachment culture in vitro, implantation in vivo, and chimera production. The result showed that ECCD1 significantly delayed embryo compaction and blastocoele formation. ECCD1treated embryos had obvious inner cell mass, positive alkaline phosphatase and pluripotent markers, normal attachment growth in vitro and implantation in vivo. In addition, the chimera production was also improved when 8cell embryos injected by embryonic stem cells were treated with ECCD1. The results indicated that compaction delay by ECCD1 antibody did not affect embryo development potential.
动物营养
Prediction of Nutrient Digestibility and Energy Concentrations Using Chemical Compositions in Meat Sheep Feeds
LIU Jie;DIAO Qiyu;ZHAO Yiguang;JIANG Chenggang;DENG Kaidong; LI Yanling;TU Yan
2012, 43(8):  1230-1238.  doi:
Abstract ( 312 )   PDF (399KB) ( 717 )  
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The objectives of this experiment were to use the data of chemical composition to develop prediction equations for nutrient digestibility and energy concentrations of meat sheep feeds. Twelve crossbred rams (Dorper♂×Smalltailed Han♀) fitted with ruminal cannulas and body weight of (47.21±1.01) kg were used in a uncompleted 12×4 Latin square arrangement. Chemical composition, nutrient digestibility and energy concentrations of each ration were determined and calculated by regression method. The results showed that, digestibility of dry matter (DM), organic matter (OM), gross energy (GE) and crude protein (CP) were positively related to their contents in diet (P<0.01), whereas they were negatively related to neutral detergent fiber (NDF) (P<0.01). Digestibility of NDF was negatively related to OM, GE and CP in diet (P<0.05), and positively related to the NDF(P<0.01). There were positive relationship between concentrations of digestible energy (DE) and metabolizable energy (ME) and the content of OM, GE and CP in diet (P<0.01), whereas they were negatively related (P<0.01) to NDF. Prediction equation of energy digestibility (ED) and ME based on chemical composition were ED(%)=194.907-0.987NDF(%)-0.901OM(%)-0.603CP(%) (R2=0.966, n=12, P<0.001) and ME(MJ·kg-1DM)=50.245-0.136NDF(%)-0.394OM(%)-0.012CP(%) (R2=0.901, n=12, P<0.001), respectively. These results indicated that there were significant relationship between nutrient digestibility, energy concentrations and chemical composition, and the equations developed in the present experiment could accurately predict nutrient digestibility and energy concentrations using chemical composition in meat sheep feeds.
Effects of Dietary Cationanion Balance on Performance of Heatstressed Beef Cattle
CAO Yufeng;;LI Qiufeng;;GAO Yanxia;;YU Haichuan;LI Jian guo;;FENG Zhihua;CHEN Fuyin;CUI Qiujia;ZHAO Huili;WANG Meimei
2012, 43(8):  1239-1246.  doi:
Abstract ( 315 )   PDF (421KB) ( 479 )  
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The objective of this trial was to study the effects of different DCAB on the production performance, blood chemistry and lymphocyte apoptosis of heatstressed beef cattle. 30 Simmental cattle with similar weight( (450±10)kg) were randomly divided into three groups,every group contains ten cattle. The diets of three experimental groups was group Ⅰ(control group, DCAB, 108 meq·kg-1 DM), group Ⅱ(228 meq·kg-1 DM ), group Ⅲ(347 meq·kg-1 DM). The results showed that, under the condition of heat stress, different DCAB value in diet didn’t influence rectal temperature, while the respiratory rate decreased linearly with increasing DCAB (P<0.05) . Compared with control group, daily weight gain of beef cattle in group Ⅱ and Ⅲ increased 29% (P<0.05) and 25% (P<0.05), respectively, feed conversion(feed/gain)decreased by 17.34% (P<0.05) and 11.46% (P>0.05), respectively. With the increasing DCAB, dietary NDF and ADF apparent digestibility improved (P>0.05), Ca, ash apparent digestibility showed quadratic change, crude protein and crude fat apparent digestibility had no significant difference. Compared with the control group, serum T4 levels of groupⅡ and group Ⅲ were increased 58.1% (P<0.05) and 31.9% (P>0.05), respectively, SOD levels increased 18.98% (P>0.05) and 48.18% (P>0.05 ), respectively. Increasing DCAB had no significant effect on serum glucose and urea nitrogen, albumin. Increased DCAB could significantly reduce lymphocytes early apoptosis (P<0.01), late apoptosis (P<0.01) and total apoptosis rate (P<0.01), reduce the proportion of lymphocytes (P<0.05) in peripheral blood of beef cattle, and improve the ability to resist heat stress. In conclusion, the suitable level of DCAB was 228 meq·kg-1 DM in diet of heatstressed beef cattle.
Effects of Forcefeeding Levels on Carcass Quality, Body Fat Deposition and Apparent Digestibility of Nutrients for Pekin Ducks
WEN Zhiguo;ZHU Yongwen;TANG Jing;XIE Ming;HUANG Wei; YU Junying;HOU Shuisheng
2012, 43(8):  1247-1254.  doi:
Abstract ( 290 )   PDF (397KB) ( 564 )  
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This experiment was conducted to determine the effects of different forcefeeding levels on carcass quality, body fat deposition and apparent digestibility of nutrients for Pekin ducks. Ninetysix 35dayold health male Pekin ducks with similar pen weight were randomly assigned into 8 treatments with 6 replicates per treatment and 2 Pekin ducks per replicate. Apparent digestibility of nutrients was determined with chromic oxide (Cr2O3) at an inclusion rate of 0.5% in basal diet. The experimental feeding period lasted 7 d from 35 to 42 d of age and the cornsoybean diet was fed to the eight groups of ducks, the control group by ad libitumfeeding and the experimental groups by forcefeeding. The forcefed intake was 260 g·d-1 at 35 d of age and increased to 300 g·d-1 at 36 d of age. The forcefed intake unchanged during the other five days and they were 300,330,360,390,420,450 and 480 g·d-1 each treatments, respectively. The results showed as follows: (1)No significant differences in dressing percentage(DP),eviscerated percentage(EP),breast muscle percentage(BMP),leg muscle weight(LMW) and leg muscle percentage(LMP)were observed among treatments(P>0.05). Breast muscle weight(BMW) increased with increasing feeding level from the control group to 390 g·d-1 (P<0.05) and then decreased (P>0.05). Based on the brokenline regression model analysis with BMW data, the optimum feeding levels for forcefeeding Pekin ducks was 390.7 g·d-1; These were significant differences among treatments in body fat deposition index(P<0.05). But body fat deposition were unchange when forcefeeding levels reached a certain value (P>0.05).(2)For the final overfeeding period, there were significant differences in apparent digestibility of dry matter and crude protein (P<0.05), whereas apparent digestibility of energy had no significant differences (P>0.05) between different treatments. In conclusion, overfeeding of Pekin ducks can induces a strong fattening of subcutaneous adipose tissues and muscles and forcefeeding has a significant effect on breast muscle growth. However, nutrients digestibility decrease and body fat deposition keep unchange with increasing forcefeeding levels.

预防兽医
Comparative Studies of Statistic Design and Analysis for Antibody Titer Kinetics after Vaccination of Chickens with Inactivated Viral Vaccines
CUI Zhizhong;ZHAO Peng;HE Hairong;ZHU Lilong
2012, 43(8):  1255-1259.  doi:
Abstract ( 284 )   PDF (464KB) ( 487 )  
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The aim of this study was to provide scientific data for establishment of better vaccination procedures in chicken breeders or layers. Different statistic designs and analysis were compared to follow dynamic changes of antibody titers to Newcastle disease virus,H5 and H9 avian influenza viruses in a chicken breeder farm of large scale. It was indicated that the statistic design and analysis by ttest for paired comparisons could prevent some false decreases in antibody titers caused by technical or systemic errors and easily detect the small but real decrease in antibody titers, such small decrease may be covered by big standard deviations due to individual variances if test for comparison between two sample means was used. The statistic design and analysis by ttest for paired comparisons made conclusions more convenient and reliable than that by ttest for comparison between two sample means in determination of real decreases in antibody titers to Newcastle disease virus, H5 and H9 avian influenza viruses.
The Two Key Sites Responsible for Drug Resistance in H3N2 Neuraminidase Gene
DONG Changying;HAN Bingxiao
2012, 43(8):  1260-1265.  doi:
Abstract ( 240 )   PDF (800KB) ( 411 )  
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To determine the key sites responsible for drug resistance in H3N2 neuraminidase gene, H3N2 wild type and three mutation type (E119D+I222L,E119D and I222L) viruses were generated by reverse genetics; The reverse genetics viruses were stable after five passages in MDCK cells; Oseltamivir resistance was detected by analyzing the kinetics of the two types of viruses in MDCK cells in the absence or in the presence of oseltamivir at the indicated times. We successfully generated H3N2 wild type and mutation type viruses; The wild type had a similar virus titer with the mutation type E119D; The virus titer of E119D+I222L and I222L type were lower than the wild type; The mutation type E119D+I222L had a high oseltamivir resistance while the wild type, E119D type and I222L type were greatly inhibited by oseltamivir. These results indicated that double mutation (E119D+I222L) type H3N2 could induce a high level of oseltamivir resistance. Thus, two amino acids in H3N2 neuraminidase gene (119, 202) are the two key sites responsible for oseltamivir resistance.
Construction and Humoral Immune Response of Recombinant Plasmid CoExpressing GP3,GP5 and M Proteins of Porcine Reproductive and Respiratory Syndrome Virus
WANG Fan;LIU Jianbin;ZHU Xiumei;LV Zhihui;MA Quanying; DU Ping;LIU XueRong;MU Kebin
2012, 43(8):  1266-1272.  doi:
Abstract ( 313 )   PDF (705KB) ( 434 )  
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The aim of this study was to construct a recombinant eukaryotic plasmid coexpressing GP3, GP5 and M proteins of porcine reproductive and respiratory syndrome virus (PRRSV). GP3, GP5 and M protein gene fragments of PRRSV LN strain were amplified by PCR and cloned into vector pcDNA3.1(+) to produce the recombinant plasmid pcDNA3.1GP3GP5M. The plasmid was transfected into COS7 cells, and immunological responses to the plasmid were investigated in BALB/c mice and piglets. The recombinant plasmid pcDNA3.1GP3GP5M was transfected into COS7 cells, PCR identification and indirect immunofluorescence assay proved that GP3, GP5M gene of PRRSV were expressed in COS7 cells. The result of Western blotting showed that the GP3 and GP5 and M proteins were coexpressed and formed fusion protein. The BALB/c mice were injected with recombinant plasmid pcDNA3.1GP3GP5M to evaluate the induced immunological responses in vivo. The specific detectable antiPRRSV neutralizing antibodies were produced in the vaccinated mice at 2 weeks and reached a peak 1:32 at 8 weeks after primary vaccination. In addition, it was observed that the 1:41:8 neutralizing antibodies of the vaccinated piglets. The results showed that the recombinant plasmid pcDNA3.1GP3GP5M has been constructed successfully, and the recombinant plasmid has well immunity, the recombinant plasmid could be a basis to eukaryotic vector vaccine for PRRSV infection.
Construction and Identification of Attenuated Salmonella typhimurium Harbouring S/N Double Fusion Genes of Porcine Transmissible Gastroenteritis Virus
HUANG Xiaobo;LI Chunsong;YANG Heng;CAO Sanjie; WEN Xintian;LIAO Xiaodan;ZHANG Xinmiao
2012, 43(8):  1273-1280.  doi:
Abstract ( 258 )   PDF (1263KB) ( 427 )  
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To provide a new vaccine for oral immunization of transmissible gastroenteritis virus (TGEV), attenuated Salmonella typhimurium harbouring S/N double fusion genes of (TGEV) was constructed and identified. The S gene fragment (2.1 kb) and the N gene fragment (1.2 kb) were respectively amplified from the recombinant plasmid 19TS and 19TN of TGEV by RTPCR, and then the two gene fragments were successively inserted into the eukaryotic expression vector pVAX1 to construction the recombinant eukaryotic expression plasmid pVAXS/N that expressing the SN double fusion gene. The plasmid pVAXS/N was identified by PCR and restrictive digestion, and then the pVAXS/N was transfected into COS7 cells through liposome transfection to identify the expressions of the two target genes by indirect immunofluorscence assay. The plasmid pVAXS/N was transformed by electroporation into attenuate S. typhimurium SL7207, the stability of pVAXS/N in SL7207 (pVAXS/N) cultured in vitro was detected. The transcription of pVAXS/N in mouse ileal tissue cells orally immunized with SL7207 (pVAXS/N), BALB/c mice were inoculated orally with SL7207 (pVAXS/N) at different dosages to detect the safety and stability in vivo. The results showed that the recombinant plasmid pVAXS/N was constructed correctly and the expression of S gene and N gene were detected in COS7 cells. The transcription and expression of S/N double fusion gene were detected in mouse ileal tissue cells after 3 days infected with SL7207 (pVAXS/N). The recombinant bacteria SL7207 (pVAXS/N) was highly stable cultured with Kanamycin Resistance medium in vitro. The recombinant SL7207 (pVAXS/N) were all safe to mouse at dosage of 0.5×109, 1×109 and 2×109 CFU. The SL7207 (pVAXS/N) were eventually eliminated from the spleen and liver at about four weeks postimmunization. These results indicated that the recombinant bacteria SL7207 (pVAXS/N) had good safety and stability, which lay a foundation for oral immunization of TGEV.
Screening of Antigen Mimotopes of Rabbit Hemorrhagic Disease Virus by Phage Display Technology
YANG Tingya;;WANG Fang;JIANG Ping;HU Bo;FAN Zhiyu;WEI Houjun;XUE Jiabin
2012, 43(8):  1281-1286.  doi:
Abstract ( 272 )   PDF (922KB) ( 524 )  
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The monoclonal antibody (McAb) A3c against rabbit hemorrhagic disease virus had been used as solidphase selective molecule to screen the epitope of RHDV by phage display technology. McAb A3c was coated on the solidphase and then three rounds of biopanning were carried out; 25 phages were selected and amplified to be identified by ELISA, and the positives were sequenced; the serum of mice immunized three times was prepared to determine the immunogenicity of the epitope. The results showed that the specific phages were enriched effectively after three rounds of biopanning; 19 of 25 phages were positive. The sequence analysis of the positive clones showed that highly homogenous sequence (GTDDMDPGTTAA) comparing to antigen was got, which is the antigen mimotopes of RHDV. The sequence DXXDP was the core amino acids in the epitope. In addition, the serum of mice reacted with antigen well and so did positive phages with hyperimmune serum of rabbit against RHDV, which indicated that the epotipe had good immunogenicity and reactionogenicity. This study provide theoretical basis for studying the epitope of RHDV and new vaccines.
Effect of Mature Chicken Interleukin-18 Protein on Its Immunoenhancement on Newcastle Disease Vaccine
ZHANG Haibin;ZHOU Mingdong;ZHOU Xia;HU Jingdong;ZHAO Hongkun;
2012, 43(8):  1287-1291.  doi:
Abstract ( 260 )   PDF (353KB) ( 378 )  
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The liposomeencapsulated mChIL18 was firstly applied on its immunoenhancement for Newcastle disease vaccine with different concentrations. In the experiment, onedayold SPF chickens were randomly divided into six groups, the blank control group, the NDV vaccine group, the NDV vaccine plus 0.2 mL mChIL18 group, the NDV vaccine plus 0.1 mL mChIL18 group, the NDV vaccine plus 0.3 mL mChIL18 group and the NDV vaccine plus vector control group.All groups were immunized on the seven days old.After that, peripheral blood was collected with anticoagulation, and the antibody level, CD4+ and CD8+ T lympholeukocytes were detected by HI and FACS at each week, respectively. Standard Virulent Strain F48E9 (100 times ELD50 per chicken) was given to the chickens at last vaccination. The results indicated that the best concentration of mChIL18 liposomes is 0.2 mL per chicken. The mChIL18 liposomes can not only enhance the cellular immune response significantly, but also enhance the humoral response, improving the protective rate of ND vaccine.
基础兽医
Dynamic Changes of Bacterial Community in Uterus and Relationship with Uterus Mucus Characteristics of Postpartum Cows
PENG Yu;WANG Yihao;ZHU Weiyun;LIU Xiuhong;HANG Suqin
2012, 43(8):  1292-1297.  doi:
Abstract ( 264 )   PDF (383KB) ( 405 )  
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Dynamic changes of predominate bacteria in the uterus of postpartum dairy cows, and the relationship of uterus mucus characterestic and dominate bacteria community were investigated. Uterus mucus of total 42 dairy cows was collected on the 10, 20, 30 and 40 days after parturition, respectively. Bacteria were isolated and subsequently identified by aerobic and anaerobic cultures using standard bacteriological techniques. The results showed that the most prevalent bacteria in uterine after parturition were E. coli (85.7%), Proteus (64.3%) and Staphylococcus sp. (61.9%). The dominant bacteria on 10, 20, 30 and 40 days after parturition were Staphylococcus sp., Clostridium perfringens, Salmonella sp., Staphylococcus sp., respectively. It was found that E. coli had high isolation rate at each timepoint. The number of bacteria isolated from white or off white mucus and yellow or earthy yellow purulent mucus (5.6±0.4,5.5±1.4) were higher than that isolated from clear mucus (3.5±1.6). The isolation rate of Salmonella, Proteus sp., Streptococcus sp. and Arcanobacterium pyogenes from white or yellow purulent mucus were more than that from clear mucus on 30 and 40 days postpartum. Clostridium perfringens was associated with yellow or earthy yellow purulent mucus on 40 days postpartum. It indicates that the dominant bacterial community changes with time after parturition, and the characteristic of uterus mucus is associated with specific pathogenic bacteria.
Florfenicol and Its Metabolite Florfenicol Amine Residue Depletion in Chicken Muscle
XIE Kaizhou;ZHANG Xiaojie;;CHEN Xuesen;XIE Xing; SUN Yingying;ZHANG Genxi;WANG Jinyu;CHEN Guohong;
2012, 43(8):  1298-1305.  doi:
Abstract ( 438 )   PDF (1000KB) ( 719 )  
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A study on florfenicol (FF) and florfenicol amine (FFA) residue depletion was conducted in chicken muscle. FF and FFA were extracted from chicken muscle with acetone and dichloromethane. The extract solution was degreased in nhexane saturated with acetonitrile, dried in nitrogen evaporator and residues were dissolved in mobile phase. FF and FFA were determinated by high performance liquid chromatography (HPLC) with fluorescence detector. The mobile phase was acetonitrilemonosodium phosphate solution(0.01 mol·L-1, containing 0.005 mol·L-1 SDS and 0.1% triethylamine)(35:65 by volume), the flow rate was 1.0 mL ·min-1. The fluorescence detection was with an excitation wavelength of 225 nm and an emission wavelength of 285 nm. Result were as follows: The limits of detection(LODs)of FF and FFA were 1.5 μg·kg-1(S/N=3) and 0.5 μg·kg-1(S/N=3), respectively. The limits of quantitation (LOQs)were 5 μg·kg-1(S/N=10)and 2 μg·kg-1(S/N=10), respectively. After the chickens orally administered successively FF capsules of 25.0 mg·kg-1, 50.0 mg·kg-1 and 100.0 mg·kg-1 of body weight one time every day for 5 days, respectively, at the first withdrawal day, the maximum residues of FF and FFA were detected. The residues were rising with the dosage and reducing with the withdrawal time. At the 7th withdrawal day, the total residues of FF and FFA were all lower than the MRLs (100 μg·kg-1); and all lower than the lowest LOD at 11th withdrawal day. The residues and total residues of FF and FFA in chicken muscle were all positively correlated with FF orally administered doses.

研究简报
Mutation Analysis and Protein Motifs Search of β1 Domain of SLADQB Gene in Rongchang Pigs
BAI Xiaoqing;LIU Wen;HUANG Wei;WANG Peng;SUN Yan;WANG Jinyong
2012, 43(8):  1306-1309.  doi:
Abstract ( 265 )   PDF (337KB) ( 503 )  
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The study was conducted to analyze mutations and search protein motifs of β1 domain of SLADQB gene in Rongchang pigs. The mutation analysis of the sequences of β1 domain of SLADQB gene from 53 Rongchang pigs was performed by multiple sequence alignment. The protein motifs of β1 domain of SLADQB gene was predicted by on line softwares. The results showed that 9 single nucleotide insertion sites and 16 single nucleotide deletion sites and 89 single nucleotide polymorphism sites were observed in 222 base pairs. 37 missense mutations were observed in 74 amino acid residues resulted by 37 SNPs. The change of amino acid type happened in 24 of 37 missense mutations. There were 174 protein motifs including 7 types in 50 amino acid sequences of β1 domain of SLADQB gene. The maximum number of protein motifs for a single sequence was 12 and the minimum number was 2. The mutations of protein motifs were observed at the 9th, 26th,45th,53th and 61th amino acid residues, which involing in 5 types of protein motifs. These results indicated that β1 domain of SLADQB gene in Rongchang pigs owned abundant genetic diversity and diversified protein motifs.
Separating Subcellular Proteins in Uncapacitation and Capacitation of Boar Sperm Cells and Identifying the Tyrosine Phosphoprotein
HU Qimeng;ZHANG Yuanyuan;CHEN Zhenliang;WANG Liangliang;LI Xinhong
2012, 43(8):  1310-1316.  doi:
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The aim of this study was to separate subsets proteins in uncapacitation and capacitation of boar sperm cells and to identify the protein tyrosine phosphorylation, and to establish the theoretical foundation for the mammalian sperm fertilization biology research.In vitro,the boar sperms were cultivated for capacitation and separated subsets proteins.The tyrosine phosphorylated proteins in uncapacitation and capacitation sperms were identified by using SDSPAGE and Western blot.In capacitation boar sperm, the degree of tyrosine phosphorylation on 126, 108, 79 ku proteins were significantly higher than them on uncapacition boar sperm. The vigor indexes and the results of SDSPAGE of uncapacitation and capacitation sperms were different,all the vigor indexes of capacitation sperms were significantly improved and these changes were related with tyrosine phosphorylation. The molecular weight of tyrosine phosphorylated proteins on membrane were approximately 25, 47 and 50 ku and on cytoplasm is about 47 ku. The degree of tyrosine phosphoproteins of 25 and 47 ku in capacitation sperm were significantly higher than them in the uncapacitation sperm(P<0.05). This phenomenon also existed in cell nucleus.In sperm nucleus,these proteins’ molecular weight were 23, 37 and 4250 ku.The degree of tyrosine phosphrylated 23 ku protein in nucleus of capacitation sperms are higher.The results suggests that the main tyrosine phosphorylated proteins were on the nucleus and membrane proteins,and a few of them are plasma proteins.
Clinical Cases Analysis of Avian Leukosis Virus Subgroup J Induced Erythroblastosis in Chicken
JIANG Yanping;WANG Yue;YU Linlin;CAI Liming;WANG Zhenzhen; XU Qingqing;WANG Guihua;CHENG Ziqiang
2012, 43(8):  1317-1323.  doi:
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Erythroblastosis was diagnosed in two flocks of Chinese local breed and one flock of layer chicken during 2010 and 2011. The age of birds was 110day, 90day and 20day, respectively. The main symptoms were characterized by emaciation, depressed and lethargy. At necropsy, numerous similar nodular lesions and resembling tumors were observed in one or more tissues. Some organs showed enlargement. A large number of erythrocytoblasts were observed in blood and bone marrow smears. Erythrocytoblasts were larger, basophilic cytoplasm and round nucleus. Histological examination showed that erythrocytoblasts were crowded in both blood vessels and mesenchym. Mitosis of erythrocytoblasts was often seen. No evidences showed Marek’s disease and reticuloendotheliosis, poisoning and metabolic diseases in these sick chickens. The PCRbased diagnosis showed that ALVJ specific fragment of 924 bp was obtained, rather than ALVA and ALVB. Immunohistochemistry results showed that a diffuse presence of ALVJ antigen in various tissues. Taken all data together, it demonstrated that erythroblastosis in the three flocks was induced by ALVJ. Erythroblastosis induced by ALVJ was found first time in China flocks. The etiology and mechanism of tumor spectrum expanded of ALVJ need to be further studied.
Expression of P216 Gene Fragment from Mycoplasma hyopneumoniae and Research on Adhesion Activity
DU Haixia;LIU Maojun;FENG Zhixin;XIONG Qiyan;BAI Fangfang;WANG Haiyan;SHAO Guoqing
2012, 43(8):  1324-1329.  doi:
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This experiment was conducted to study the adhesion activity of P216 protein and establish the model of adhesion protein of Mycoplasma hyopneumoniae (Mhp). According to analysis, the fragment of P216 gene with hydrophilicity, antigenicity and good adhesion was chose. P216 gene fragment was amplified by PCR from Mhp NJ strain and inserted into expression vector pET32a(+), and the recombinant plasmid pET32a(+)/P216 was constructed. After IPTG induction, the immunological and adhesion activity of the recombined protein was detected by Western blot and indirect immunofluorescence assay. The results showed that, the PCR product of the target gene was l 636 bp, the molecular weight of recombinant protein was 80.1 kDa by SDSPAGE, and Western blot results showed that recombinant protein had satisfactory immunogenicity. Indirect immunofluorescence assay showed that the recombinant protein could produce occupied inhibition to the Mhp adhering with SJPL cells. These results indicated that the P216 protein had good adhesion activity, and could adhere SJPL cells. It provides new ideas for research of the other adhesions from Mycoplasma hyopneumoniae.
研究简报
The Study of Prokaryotic Expression and Immunogenicity of E Gene of Japanese Encephalitis Virus Guizhou Isolation Strain
TANG Deyuan;WANG Feng;MA Ping;LUO Xianfeng; LI Chunyan;ZENG Zhiyong;XU Jian;LIU Jian
2012, 43(8):  1330-1336.  doi:
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Japanese Encephalitis Virus (JEV) was isolated from a porcines intumescent testis. After specimens were made into homogenate and filtrated,it was inoculated to passage cells and injected into rat brain to isolate JEV, then JEV was identified by RTPCR. According to published cDNA sequence of E gene in GenBank of JEV SA14 and SA14142 strain, the synthesized specific primers were designed. E gene fragments of JEV Guizhou strain were amplified by using RTPCR. The PCR products were sequenced and comparatively analyzed. The products of RTPCR were cloned into pMD19T vector to form recombinants pMD19TE, then were subcloned into Prokaryotic Expressing Vector pET32a (+) after double enzyme digestion and sequence analysis. The pET32aE was transformed into the E. coli strain BL21 (DE3) and the expression of the E protein was induced. After analysis by SDSPAGE electrophoresis of the induced expressed protein and Western blot, we got about 59 kDa expressed protein in line with the expected size. Through further extracting and purifying the target protein, the expressed protein was used to immunize mice by adding adjuvant, and then the antibody titers of mice were detected. The results showed that the target protein induced antiJE antibody in mice. This study laid the foundation of the research of subunit vaccine of Japanese Encephalitis.