检测中国对虾非包涵体型杆状病毒的PCR方法的建立

中国对虾非体型杆状病毒（ＰｃＮＯＢＶ）是中国大陆养殖的中国对虾（Ｐｅｎａｅｕｓｃｈｉｎｅｎｓｉｓ）暴发性流行病－－白斑征的主要病原,与台湾流行的ＰｍＮＯＢⅢ、泰国的ＰｍＮＯＢⅡ及日本的ＲＶ－ＰＪ（＝ＰＲＤＶ）有相似的致病特征、形态学及组织病理学特征。为了角ＰｃＮＯＢＶ与ＰｍＮＯＢⅢ两种地域分布邻近的毒株基因水平的关系,并建立早期、敏感、特异的检测技术,利用氯化铯密度梯度超速离心技术分离纯化了ＰｃＮ     

应用双色荧光原位杂交技术检测克氏综合征

探讨用双色荧光原位杂交技术（dual-color fluorescence in situ hybridization,D-FISH）检测性染色体数目异常克氏综合征的应用价值,建立常规分裂期染色体和间期细胞FISH技术的实验方法。以Biotin标记的X染色体α-卫星DNA（pBamX7）探针和以Digoxigenin标记的Y染色体长臂末端重复序列（pY3.4）探针对19例克氏综合征标本同时进行外周血染色体及其间期细胞核的原位杂交,分别用Avidin-FITC和Rhodamine-FITC及其Anti-avidin进行信号的检测与放大,DAPI复染。于Olympus AX-70型荧光显微镜下,分别通过WIB、WIG及其WU滤光镜观察杂交信号及其染色体或间期核背景,并统计外周血中期染色体及其间期细胞核的杂交信号颗粒数量。在显微镜下可见以Biotin标记的pBamX7探针显示2个绿色杂交信号,以Digoxigenin标记的pY3.4探针显示1个红色杂交信号,染色体或间期核背景经DAPI复染显示蓝色;18例出现XXY杂交信号的细胞,染色体及其间期细胞核杂交平均出现率分别为95.89%和95%,明显大于正常对照标准值2.75%,证实核型为47,XXY,与染色体检测的结果一致;其余1例染色体核型检测为嵌合体,XXY杂交信号细胞出现率为92%,同时检出6.7%的XY杂交信号细胞（>正常对照标准值4.17%）。用FISH 技术检测性染色体数目异常克氏综合征具有快速、敏感度高、信号强、背景低、多色等优点,故FISH 技术在产前诊断检测领域中显示其重要的应用价值和发展前景。 Abstract:The objective of the work is to study the technique of dual-color fluorescence in situ hybridization(D-FISH) and its application value in the diagnosis of sex chromosomal count abnormality Klinefelter syndrome and establish an experimental approach to metaphase chromosome and interphase nucleus FISH technique.Biotin labeled alpha satellite X-chromosome DNA(pBamX7) probe and Digoxigenin labeled Y-chromosome long arm terminal repetitive sequence (pY3.4) probe were hybridized with pre-treated slides of peripheral blood chromosome and interphase nucleus in 19 cases of Klinefelter syndrome specimens.After being washed,the slides were treated with Avidin-FITC,Rhodamine-FITC and Anti-avidin,amplified with an additional layer and counter-stained with DAPI in an antifade solution.The hybridization signals,chromosomal or interphase nucleus settings were observed respectively with WIB,WIG and WU filters under fluorescence microscope Olympus AX-70,and the number of metaphase chromosome and interphase nucleus in the peripheral blood was counted.It was observed under the microscope that the Biotin labeled pBamX7 probe showed 2 green hybridization signals and that the Digoxigenin labeled pY3.4 probe showed 1 red hybridization signal.Chromosome or interphase nucleus counter-stained with DAPI showed blue.The average signal rate of chromosome and interphase nucleus hybridization was 95.89% and 95% respectively,significantly higher than the normal control (2.75%).Karyotype 47,XXY was confirmed,which agrees with the chromosomal findings.One case showed mosaic nuclei.XXY chromosome hybridization signal rate was 92% and XY hybridization signal rate was 6.7%,higher than the normal control rate of 4.17%.FISH is a valuable technique in diagnosing sex chromosomal count abnormality Klinefelter syndrome with the merits of fast speed,high sensitivity,strong signal,low background and multiple color.Therefore,FISH technique can find wide application and potential in prenatal diagnosis.     

枯草芽孢杆菌BSD-2对黄瓜叶际微生物数量及菌群结构的影响

【目的】阐明枯草芽孢杆菌BSD-2菌株抗菌素、芽孢及发酵液不同施入形式对黄瓜叶际微生物数量及菌群多样性的影响,为该菌株的应用提供理论基础和依据。【方法】以黄瓜为实验材料于室内进行盆栽实验,设置于接种灰霉病原菌前后喷施抗菌素、芽孢、发酵液,单独接种灰霉病原菌和无菌水空白对照,共8个处理。采用传统微生物分离的方法,利用PB、高氏1号和PDA培养基分离并统计叶际可培养细菌、放线菌和真菌的数量;采用Bio-Rad公司的DcodeTM System进行样品的DGGE分析,对条带数量和灰度进行定量分析,采用UPGMA法对各条带进行聚类分析;采用Shannon-Wiener指数、Pielou均匀度指数和丰富度分析可培养微生物和DGGE图谱中的群落多样性;并将条带回收测序。【结果】枯草芽孢杆菌BSD-2抗菌素、芽孢及发酵液以接种灰霉病原菌前喷施的效果最佳,均可提高可培养细菌的数量,降低可培养真菌的数量;可使叶际微生物的Shannon-Wiener多样性指数、Pielou均匀度指数和丰富度指数增加;其中抗菌素处理对叶际微生物种群结构的影响最小。【结论】枯草芽孢杆菌BSD-2的发酵液可有效抑制真菌的数量;芽孢可有效提高细菌及放线菌的数量;抗菌素则利于保持叶际微生物的丰富度及多样性指数。     

抗体介导的慢性排斥反应中内皮细胞基因的变化及分子机制

目前,抗体介导的慢性肾移植排斥反应难以预测,尚无有效的治疗方法。研究表明,通过对内皮细胞相关基因的分析,可以提高抗体介导的慢性肾移植排斥反应术前及术后的诊断特异性。该病的发病机制主要包括四个方面：（1）血小板在内皮细胞附近聚集凝固能力增强;（2）内皮细胞趋化炎性细胞作用增强;（3）干扰素-γ（IFN-γ）杀伤作用增强：（4）内皮修复再生能力降低。基因分析结合抗体分析有助于深化理解抗体介导的肾移植排斥反应的发病机理、提高诊疗效果、为新药开发提供新靶点,使肾移植成功率大幅提高成为可能。该篇综述将就上述内容作一介绍。