玉米食醋主要是以玉米、高粱为主要原料经发酵而成的具有多种功能的酸性调味品.随着传统食醋生产工艺的不断改善,安全水平不断提高,但仍存在一些潜在的危害因子.文章主要探讨了玉米化学成分变化对玉米赤霉烯酮提取的影响,进而有效提高ELISA方法检测玉米赤霉烯酮的稳定性,实验以重新审视样品前处理这一最基础的科学问题,在样品前处理阶段选取经过180 s粉碎的空白玉米为实验样品,以玉米赤霉烯酮为研究对象,采用ELISA法和UPLC-MS法联合检测,以测定玉米赤霉烯酮提取率为指标,分析玉米样品中在不同放置环境下、放置不同时间段的粉碎玉米样品中水分、粗脂肪、粗蛋白、灰分等化学成分的变化与玉米赤霉烯酮添加回收率的相关性.结果表明,不同储存环境随着储存时间延长,水分含量、粗脂肪含量与玉米赤霉烯酮的添加回收率具有一定的相关性,水分影响大于粗脂肪.粗蛋白、灰分与玉米赤霉烯酮的添加回收率无明显相关性.

选取了黑熊的9个微卫星基因座(ABB1、ABB3、ABB4、ABB5、ABB6、ABB7、ABB10、ABB11、ABB12)进行遗传多样性研究.结果:东北黑熊养殖群体的9个微卫星座位共检测到60个等位基因,平均为6.67个;9个微卫星基因座的平均多态信息含量为0.589 7,其中6个微卫星基因座的多态信息含量(PIC) ＞0.5,9个微卫星基因座的平均观察杂合度、平均期望杂合度分别为0.536 3、0.625 2.由此证明,9个微卫星标记均具有高度多态性,可用于东北黑熊的遗传多样性分析.

The research aimed to discuss the feasibility of make the genegic diversity research by using U.t.ussuricus hair wholly as tested materials.With four adult U.t.ussuricus of Yanbian Baitou mountain pharmacectic company,co,ltd bear breeding factory as tested materials,50 and 100 hairs were picked up five times in two bear cages and 3 mL whole blood were taken out and 1g subcutaneous connective tissue were cut to make DNA extraction in different methods.The blood and subcutaneous connective tissue were obtained,when the two bears were narcotized.With microsatellite primers ABB12 and UT35 as primers,3template concentration including 20,50,100 ng.μL-1 were set up to make PCR amplification.Using 50 ng.μL-1 as template concentration could obtain better PCR products,which had better resolving effects on the amplification genes in polyacrylamide gel electrophoresis.The quantity and quality of PCR products among hair,blood and tissue DNA samples had no obvious difference.The hair number had suitable correlation with the concentration of DNA.When the PCR products of 15 whole-hair samples were used in the native polyacrylamide gel electrophoresis,clear bands could be observed.The research results further proved that DNA from whole-hair samples could be taken as the materials for U.t.ussuricus genetic diversity research.

为了筛选得到免疫效果较理想的核酸疫苗,采用pVAXⅠ为真核表达载体,新孢子虫P0基因为目的基因,构建了重组质粒pVAXⅠ -NcP0.用重组质粒免疫成年沙鼠60只,剂量为100 μg/只,每隔14 d免疫1次,共免3次,首免后每周尾静脉采血1次,三免后第2周,每组处死7只沙鼠,无菌取脾.采用间接荧光抗体试验检测沙鼠血清抗体效价,流式细胞仪检测脾T淋巴细胞亚群CD4+、CD8+数量.结果:Cp-M佐剂组、pVAX Ⅰ -NcP0组的抗体效价均高于pVAXⅠ 空载体组和对照组,其中Cp-M组效价最高；脾T淋巴细胞亚群数量测定发现,疫苗Cp-M佐剂组、pVAXⅠ -NcP0组的CD4+细胞含量均高于pVAXⅠ空载体组和对照组,Cp-M组、pVAXⅠ -NcP0组与pVAXⅠ空载体组和对照组均差异极显著(P＜0.01),Cp-M组、pVAXⅠ -NcP0组的CD8+细胞含量均高于pVAX Ⅰ空载体组和对照组,Cp-M佐剂组与pVAXⅠ空载体组和对照组差异显著(P＜0.05).表明DNA疫苗pVAXⅠ -NcP0既能较好地刺激细胞免疫又能产生一定的体液免疫反应,可作为候选核酸疫苗进行下一步本体动物免疫试验研究.

In order to establish the method for detecting GPV,research rabbit anti-GPV IgG was used to be capture antibody and anti-GPV monoclonal antibody was used to be detection of antibody.Based on chess-board experiment, the optimal coating concentration of rabbit anti-GPV IgG is 16 mg/L,the optimal working concentration of anti-GPV monoclonal antibody is 10.8 mg/L,The optimization of dilution for enzyme-labeled second antibody is 1:4 000,and double sandwich ELISA whose sensitivity of detecting GPV is 0.312 mg/L.was established.Dead gooses in a goose ranch in Yanbian were detected by this method.The positive rate is 75.79%.The concordant rate with virus neutralization test is 91.11%.Double sandwich ELISA established by this research provides a convenient and fast sero-diagnosis for detecting GPV and epidemiologic survey in district.

In order to validate the immunological protection effect of two different inactivated vaccines against Neosporiasis tachyzoites.The gerbils were vaccinated twice with two different adjuvant vaccine.21 days later with tachyzoites(1×106 Per gerbils) subcutaneous infection and observation gerbils information for death.The results showed that the protection rate is 85%(17/20)of the first kind vaccine,the other(Ⅱ)vaccine protection rate 80%(16/20).Amplification Neosporiasis specif fragment from the death gerbils tissue DNA.

为了研究鹅细小病毒灭活苗、亚单位疫苗、核酸疫苗3种疫苗对Balb/c小鼠的免疫效果,试验自制了这3种疫苗,对Balb/c小鼠进行3次免疫,应用间接ELISA方法测定体液免疫水平。结果表明:用小鹅瘟灭活苗、小鹅瘟VP3基因亚单位疫苗免疫Balb/c小鼠,在三免后第21天抗体的P/N值达到峰值,分别为8.21,3.56;小鹅瘟VP3基因核酸疫苗在三免后第28天,抗体的P/N值达到峰值,为2.09。说明这3种疫苗中小鹅瘟灭活苗对Balb/c小鼠的免疫效果最好。

Full length gene fragment of Neospora caninum ribosomal phosphoprotein(NcP0) was amplified by PCR,and then cloned into the eukaryotic expression vector pVAX1,thus constructing pVAX1-NcP0.The recombinant vector was transfected into Vero cell by lipidosome method.RT-PCR and immunofluoescence assay(IFA) were performed to determine the transcription of the target gene.The results showed that the cloned full length gene was 1 379 bp;RT-PCR demonstrated that the gene has been successfully transcribed;IFA test indicated that the gene was successfully expressed.In the study,the NcP0 eukaryotic expression vector was successfully constructed,and after the vector was transfected into Vero cells,NcP0 obtained the transient expression.

新孢子虫病(Neosporosis)是由犬新孢子虫(Neospora caninum)或类新孢子虫(Neosporalike)寄生于宿主动物所引起的多种家畜共患的一种原虫病[1].它可引起孕畜流产或死胎以及新生儿的运动障碍和神经系统疾病[2].目前,国外已研制出ELISA诊断试剂盒,但价格昂贵,不适合临床推广应用.而国内虽然建立了多种血清学诊断方法,但所用抗原均为重组蛋白,其敏感性、特异性将会受到不同程度的影响.因此,针对上述问题,试验对新孢子虫虫体抗原进行了分析,并应用新孢子虫虫特异性抗原建立了特异、敏感、稳定的血清学诊断方法,旨在为今后新孢子虫病的诊断和预防研究奠定基础.