牛瑟氏泰勒虫p23-IL-18融合基因的原核表达     

白杨  许应天 《中国兽药杂志》2014,48(7):6-9

为了研发牛瑟氏泰勒虫病基因工程亚单位疫苗,本实验将p23-IL-18融合基因克隆到pET-28a原核表达载体,构建pET-28a-p23-IL-18重组原核表达质粒,经BamHⅠ和EcoRⅠ双酶切鉴定后在37 ℃用IPTG诱导表达,对表达产物进行SDS-PAGE和Western blotting分析。结果表明,成功构建牛瑟氏泰勒虫pET-28a-p23-IL-18原核表达质粒,目的基因在大肠杆菌中成功获得表达,融合蛋白的分子量约为48ku,并被牛瑟氏泰勒虫阳性血清所识别,具有良好的反应原性。本研究结果为牛瑟氏泰勒虫病基因工程亚单位疫苗的制备奠定了基础。  相似文献   

隐孢子虫不同基因型P23基因的克隆及序列比较   总被引：2，自引：1，他引：1  

胡义彬  米荣升  陈兆国 《动物医学进展》2010,31(12)

为克隆隐孢子虫不同基因型子孢子表面抗原P23基因,比较其序列差异,提取上海地区分离的隐孢子虫鼠基因型(Cryptosporidiummouse genotype)、隐孢子虫兔基因型(Cryptosporidiumrabbit geno-type)、隐孢子虫猪基因型Ⅱ(Cryptosporidiumpig genotypeⅡ)总RNA,经RT-PCR扩增P23基因,克隆到pMD18-T载体中,进行序列测定,并与GenBank上下载的微小隐孢子虫(Cryptosporidium parvum)序列进行同源性比对。结果显示,从隐孢子虫3个基因型中均扩增出了P23基因。与微小隐孢子虫P23基因核苷酸序列比较,隐孢子虫鼠基因型、兔基因型、猪基因型ⅡP23基因同源性分别为97.6%、97.3%、97.3%,氨基酸序列同源性分别为97.3%、97.3%和96.4%。获得了隐孢子虫鼠基因型、兔基因型、猪基因型Ⅱ子孢子表面抗原P23基因。  相似文献   

nm23基因的表达与皮肤鳞癌发生、发展及转移的关系     

李慧忠  李顺凡  吴国凤  樊翌明  吴志华 《湛江医学院学报》2003,21(5):440-441,444

目的：观察nm23表达与皮肤鳞状细胞癌(鳞癌)发生、发展与转移的关系。方法；采用免疫组织化学及图像分析技术检测66例皮肤鳞癌、12例淋巴结转移灶、21例假上皮瘤样增生及20例正常皮肤中nm23表达情况。结果：皮肤鳞癌中nm23表达下调，但鳞癌中nm23表达与鳞癌的转移、TNM分期、病理分化程度及发病部位无关。结论：nm23基因表达下调是发生在鳞癌演进过程中的一个早期事件。nm23可能不具有抑制皮肤鳞癌转移的功能。  相似文献   

水稻新品种"延粳23号"的选育     

玄英实  元东林  程正海  杭秋瑜 《延边大学农学学报》2007,29(2):98-100,124

“延粳23号”是从韩国引进的杂交后代材料中选育出来的中晚熟水稻新品种,具有高产、优质、耐冷、适应性广等特点,2001年通过全国农作物品种委员会审定.  相似文献   

NCSU23、SOF及氨基酸对猪孤雌激活胚胎体外发育的影响*     

吴中红  邢凤英  刘国世  曾申明  朱士恩  张忠诚 《农业生物技术学报》2003,11(5):511-515

研究了培养基(NCSU23和SOF)和添加氨基酸对猪孤雌激活胚胎体外发育的影响，以探明猪早期胚胎体外培养的最佳体系。实验结果表明，在高氧(5％CO2的空气)或低氧环境(5％CO2：7％O2：88％N2)中，猪的孤雌发育胚胎培养在添加氨基酸的合成输卵管液(SOFaa)中其囊胚发育率显著低于NCSU23培养液；但培养液中添加氨基酸能显著提高卵裂率，培养后期去除氨基酸可减小氨基酸造成的不良影响。胚胎培养早期(前两天)添加氨基酸，不能提高囊胚发育率和囊胚细胞数。  相似文献   

干旱胁迫下不同品种小麦4种功能基因的表达模式及相关生理指标分析     

韩翠英  刘秉焱  刘虎岐 《西北农业学报》2015,24(10):28-34

采用实时荧光定量PCR(Real-time PCR)对干旱胁迫下脂质转移蛋白基因(TaLTP1)、膨胀素基因(TaEXPB23)、水通道蛋白基因(TaAQP7)和果聚糖6-果糖基转移酶基因(Ta6-SFT)在4种小麦叶片中的表达情况进行分析;通过测定相对水分质量分数和果聚糖质量分数来判断小麦生长发育状况。结果显示:干旱胁迫48h内,4种基因在不同抗旱性小麦叶片中的表达模式不同,干旱敏感型小麦中4种基因的相对表达量先上升后下降,恢复到正常水平;干旱耐受型小麦中4种基因的相对表达量先上升后下降,但仍维持较高表达水平;干旱耐受型小麦具有较高的相对水分质量分数和果聚糖累积量。以上结果表明,4种基因参与小麦干旱胁迫应答,4种基因的表达模式可以作为鉴定小麦抗旱的分子指标。本研究可为准确快速地鉴定小麦品种抗旱性提供重要分子依据。  相似文献   

微小隐孢子虫P23基因在毕赤酵母中的表达及初步应用     

黄燕  米荣升  周鹏  曹薇  杨晓娇  王向佩  王晓娟  石凯  陈兆国 《中国动物传染病学报》2012,20(6):56-62

为将微小隐孢子虫(Cryptosporidium parvum)P23基因在巴斯德毕赤酵母(Pichia pastoris)系统中进行表达,利用表达蛋白初步建立隐孢子虫病间接ELISA诊断技术,设计引物从微小隐孢子虫基因组DNA中扩增P23基因序列,构建pPIC9K-P23重组质粒,在毕赤酵母中进行表达,用阴离子交换层析柱进行纯化。以重组P23纯化蛋白为抗原建立间接ELISA检测方法,对现场采集的猪血清样品进行检测。SDS-PAGE显示所表达的蛋白大小约为23 kDa。Western blot检测表明该蛋白能与兔抗P23蛋白血清特异性结合。用建立的间接ELISA技术对186份猪血清样品进行检测,阳性率为83.3%。本研究获得了真核表达的P23重组蛋白,初步建立了微小隐孢子虫病间接ELISA诊断技术,为隐孢子虫病的诊断和流行病学调查打下了基础。  相似文献   

微小隐孢子虫CP15-P23-CP15/60基因的融合表达及抗血清的制备     

秦培兰  米荣升  黄燕  周鹏  张国恩  苏庆美  呼高伟  曹薇  陈兆国 《动物医学进展》2012,33(7):54-59

以微小隐孢子虫基因组DNA为模板,PCR扩增获得子孢子表面抗原CP15、P23和CP15/60基因。利用重叠延伸PCR(SOE PCR)将该3段基因片段串联在一起,各基因片段之间引入柔性氨基酸接头(GGGGS)编码基因。将串联基因克隆到原核表达载体pET-28a(+)上,构建重组表达载体,转化到大肠埃希菌BL21(DE3)中进行诱导表达,将纯化的重组蛋白免疫Balb/c小鼠制备多克隆抗体。结果表明,获得了CP15-P23-CP15/60融合基因,并在大肠埃希菌中高效表达,Western blot显示重组蛋白能被牛抗微小隐孢子虫阳性血清识别,制备的多克隆抗体能被重组蛋白特异性识别,表明获得的重组蛋白具有较好的抗原性。  相似文献   

仔猪常见12种致病菌的23S rRNA基因序列分析     

伍诚意  陈小玲  王小莺  梁之昶  相磊  章振华  季海峰 《黑龙江畜牧兽医》2008,(11)

以107条从NCBI下载和测序的12种仔猪常见致病菌的23S rRNA基因序列为研究对象,运用DNASTAR软件进行系统发育分析.结果显示:以各种细菌的23S rRNA基因代表序列所进行的种间序列分析结果与伯杰氏细菌分类手册和http://www.wiki.cn/wiki/细菌分类表的分类结果一致,也与这12种菌的16S rRNA基因序列分类结果一致. 因此,在23S rRNA基因序列保守区设计通用引物,在其变异区设计各种细菌特异性探针,用PCR捕捉被检样品中未知细菌的23S rDNA片段并与种特异性23S rRNA基因探针进行杂交,有可能将未知细菌鉴定到种.  相似文献   

Change of intestinal flora and its relationship with IL-23/IL-17 axis in patients with ulcerative colitis     

MA Xu-yuan  DAI Zhi-feng  WANG Hui-chao  YANG Jing-nan  TANG Xiang-yang  KANG Yu-hua  DING Chun-sheng  LI Yu-xia  YANG Rui-lin  LIN Xu-hong 《园艺学报》2018,34(5):884-892

AIM:To investigate the change of intestinal flora distribution and its relationship with interleukin-23 (IL-23)/IL-17 axis in ulcerative colitis (UC) patients. METHODS:The fresh fecal samples from 20 patients with active UC and 20 healthy controls were collected. The distribution of the flora was analyzed by direct smear and traditional bacterial culture. The changes of bacteria were detected by real-time PCR. The hemoglobin, albumin, erythrocyte sedimentation, and C-reactive protein levels were tested routinely. Both normal and damaged mucosal tissues of UC patients were examined and obtained by colonoscopy, and further assessed by Mayo scoring, Baron grading and HE staining. The expression of IL-17 and IL-23 was observed by immunohistochemistry and Western blot. RESULTS:(1) The degree of flora imbalance in active UC patients was higher than that in the healthy controls (P<0.05). (2) The results of aerobic culture showed that the number of Escherichia coli in the UC patients was significantly lower than that in the normal controls (P<0.01), while Enterococcus was increased obviously (P<0.01). The results of anaerobic culture revealed that the numbers of Bacteroidetes, Bifidobacterium bifidum and Lactobacilli in the UC patients were significantly decreased (P<0.01). (3) Quantitative analysis of target bacteria showed that the relative quantification of Escherichia coli, Bacteroidetes, Bifidobacterium bifidum and Lactobacilli in the UC patients was significantly lower than that in the normal subjects, and the number of Enterococcus was significantly increased (P<0.01). (4) Compared with control group, no significant change of hemoglobin in the UC patients was ovserved, albumin was significantly decreased (P<0.05), but erythrocyte sedimentation and C-reactive protein levels were elevated obviously (P<0.01). (5) The Mayo score, Baron grade, and histopathological score were all increased (P<0.01). (6) High IL-17 and IL-23 expression levels were detected in the UC patients (P<0.01). (7) Correlation analysis showed that the average absorbance values of IL-17 and IL-23 expression were positively correlated with Baron grade (r=0.717, P=0.02; r=0.849, P=0.016) and pathological score (r=0.660, P=0.03; r=0.675, P=0.032). Meanwhile, the average absorbance value of IL-23 expression was negatively correlated with the number of Escherichia coli (r=-0.699, P=0.025), and positively correlated with Enterococcus (r=0.872, P=0.010). Furthermore, the average absorbance value of IL-17 expression was positively correlated with Enterococcus (r=0.764, P=0.046), and both of them were not correlated with other bacteria. CONCLUSION:Obvious flora imbalance exists in active UC patients, changed intestinal microflora is closely related with the degree of inflammation. IL-23/IL-17 axis, as a key factor in the development of UC, may be related to the changes of intestinal microflora. The interaction between intestinal microflora and IL-23/IL-17 axis plays an important role in the pathogenesis of UC.  相似文献