转基因玉米转化体特异性寡核苷酸芯片测试方法的研制

根据7种转基因玉米Bt11、Bt176、Mon810、Mon863、TC1507、GA21和NK603的重组DNA结构,利用Primer premier 5.0引物设计软件分别设计转化体特异性引物,优化PCR反应条件,对引物特异性及灵敏度进行验证。结果表明,所设计引物特异性强,灵敏度达0.1%。在植物基因组与外源基因的连接区域,利用Primer premier 5.0和Oligo 6.0软件分别设计和筛选7种转基因玉米40mer oligo转化体特异性探针,制备转基因玉米的寡核苷酸芯片,并对芯片检测的特异性、重复性及检测灵敏度进行检验。证明所设计探针特异性强,灵敏度达0.01%。与PCR检测方法相比,基因芯片检测法灵敏度高、特异性强,可有效检测及鉴定多种转基因玉米,大大提高检测的准确率和效率。     

稻谷散装粮堆储粮害虫两种检测方法效果的比较

应用粮虫陷阱检测器和常规风力扦样器对高大平房仓中的仓虫进行监测,结果袁明,粮虫陷阱检测器比风力扦样器应用简便、省工省力,且能捕到更多种类和数量的害虫.经二者测虫比较,探讨了用粮虫陷阱检测器监测虫情的等级标准.     

TaqMan探针荧光RT-PCR检测狂犬病病毒

根据GenBank已公布的狂犬病病毒(rabies virus,RV)核蛋白(N)基因序列设计并合成一对特异性的引物和探针,建立基于TaqMan探针的荧光RT-PCR检测狂犬病病毒方法。对狂犬病疫苗提取核酸后进行RT-PCR扩增,将目的条带切胶回收,克隆测序,重组质粒作为标准阳性对照。对建立的TaqMan探针荧光RT-PCR方法做灵敏度、特异性、重复性及稳定性试验。结果显示,该方法可以达到10拷贝/μL的灵敏度,可以将狂犬病病毒与犬瘟热病毒、犬细小病毒、犬腺病毒、犬冠状病毒和犬副流感病毒分开,方法重复性好,稳定可靠。     

Establishment of Digital RT-PCR Assay for Detection of Swine Influenza Virus H3N2 Subtype

A new assay for the detection of swine influenza virus (SIV) was developed with a novel nucleic acid probe——Molecular beacon in this study. The specific primers and molecular beacon probes were designed according to the conserved region of H3 and N2 genes of SIV H3N2 subtype. A digital RT-PCR assay was developed for detection of SIV H3N2 subtype. The results showed that SIV H3N2 subtype could be identified simultaneously on this microarry with high sensitivity and reproducibility,which could reach to 10⁶ dilute viruse. The conclusion was that the digital RT-PCR method could analyze quantitatively the RNA templates.On the identification of H3N2 SIV,the digital RT-PCR method was much more scientific than Real-time quantitative PCR method.     

H1、H3亚型猪流感病毒双重实时荧光定量RT-PCR检测方法的建立及应用

为建立一种快速、准确地检测H1和H3亚型猪流感病毒（swine influence virus,SIV）的方法,根据H1和H3亚型SIV血凝素（hemagglutinin,HA）基因保守序列,分别设计2对特异性引物和2条TaqMan探针,建立双重实时荧光定量RT-PCR方法。结果显示,该方法敏感性高,可检测到最低拷贝数为10²拷贝/μL;重复性良好,重复孔Ct值的变异系数均在5%以下;特异性好,除H1和H3亚型SIV外,H4、H5、H7、H9亚型SIV、猪瘟病毒、猪繁殖与呼吸综合征病毒、口蹄疫病毒、伪狂犬病病毒检测均为阴性;在田间样品中成功检出1株H1和1株H3亚型SIV,检出率为1.16%。该方法准确、快速、灵敏、特异,可为H1、H3亚型SIV的快速鉴别检测及流行病学调查提供有效的技术手段。     

大洋臀纹粉蚧和南洋臀纹粉蚧TaqMan实时荧光PCR检测方法

臀纹粉蚧属有多个种类是重要的农业害虫,大洋臀纹粉蚧(Planococcus minor(Maskell))和南洋臀纹粉蚧(Planococcus lilacius Cockerell)是我国有重要检疫意义的有害生物.这两种臀纹粉蚧经常从进口泰国和东南亚水果口岸检疫中截获,但形态学方法很难进行准确鉴定.本研究首次利用mtDNA COI基因设计了两条特异性探针,应用TaqMan实时荧光PCR方法对大洋臀纹粉蚧和南洋臀纹粉蚧进行了快速准确鉴定.     

Embedded terminal receiving capability matching in multimedia file transmission

A novel transport protocol for large scaled multimedia file EMFTP (Embedded Multimedia File Transfer Protocol) is proposed to match the receiving capability of embedded terminal with sufficient network bandwidth. EMFTP is an application level protocol using UDP, and adopts a method of uniform retransmission as its error recovery strategy and it also introduces an improved DAIMD (AIMD with decreasing increases) rate control algorithm based on bandwidth probe. It can improve the data transfer rate with a low resource taken of embedded client. As the experimental result shows, compared with UDT and FTP protocol implemented on embedded terminal, EMFTP data transfer rate is increased by 3% and 17% respectively, the embedded client CPU occupancy ratios are reduced by 50.3% and 42.3% separately, and memory usage reduced by 15% and 4%.     

土壤中南方根结线虫的实时荧光PCR检测和定量

为了准确测定土壤中南方根结线虫Meloidogyne incognita的群体密度,根据序列特异性扩增区标记设计了南方根结线虫特异性引物和TaqMan探针,分别建立了卵和2龄幼虫的实时荧光PCR定量检测体系,并将该检测体系与显微镜计数法进行了比较.结果表明,引物Mi-F/R及探针Mi-probe对南方根结线虫具有高度特异性,建立的标准曲线循环阈值与卵和2龄幼虫数量的对数值之间具有良好的线性关系,相关系数分别为0.9771和0.9853,扩增效率均为90％,检测灵敏度为10-1个卵及10-3条2龄幼虫;对10个田间土壤样本的检测显示,实时荧光PCR定量检测与显微镜计数法测定的南方根结线虫卵和2龄幼虫数量呈显著正相关,且两种方法对2龄幼虫的测定结果无显著差异.     

厌氧去除活性污泥中痕量有机污染物:15年实验研究

Knowledge of cellular metal homeostasis will provide a better understanding of the mechanisms involved in metal tolerance and hyperaccumulation in metal-hyperaccumulating plants. Energy dispersive X-ray spectrometry (EDS) was used to determine the localization of cadmium (Cd) in leaves of the Zn/Cd hyperaccumulator Picris divaricata which had a shoot Cd concentration of 565 mg kg 1 after 2 weeks of growth in solution culture supplying 10 μmol L-1 CdCl2 . The results indicated that Cd was distributed mainly in the trichomes, upper and lower epidermis and bundle sheath cells, with a relatively low level of Cd in mesophyll cells. Mesophyll protoplasts isolated from leaves remained viable after 24 h exposure to CdCl2 at a concentration up to 1 mmol L-1 , indicating their high tolerance to Cd. The intracellular Cd was visualized by staining with Leadmium Green dye, a cellular permeable Cd fluorescence probe. The results showed that the majority of protoplasts (> 82%) did not accumulate Cd, with only a minority (< 18%) showing Cd accumulation. In the Cd-accumulating protoplasts, Cd accumulation was depressed by the addition of Fe2+ , Mn2+ and the metabolic inhibitor carbonyl cyanide m-chlorophenylhydrazone (CCCP), but not by Ca 2+ or Zn2+ . Furthermore, the entire process of Cd uptake from external solution into the cytoplasm and subsequent sequestration into vacuoles was successfully recorded by confocal images. These results suggested that reduced cellular Cd accumulation and efficient Cd vacuolar sequestration in mesophyll cells might be responsible for cellular Cd tolerance and distribution in the leaves of P. divaricata.     

辽东栎不同方位边材液流季节动态及其对蒸腾耗水测算的影响

利用Granier树干液流测定系统,对黄土高原半干旱区天然辽东栎树干不同方位的边材液流动态进行了长期监测,并同步监测环境因子(空气温度、湿度、太阳辐射等),分析了不同方位液流通量密度的相互关系、对液流通量和蒸腾耗水量测算的影响以及与环境因子的关系。结果显示,辽东栎东、西、南、北4个方位测得的液流通量密度存在显著差异,并显著相关。辽东栎生长季内(5-10月)单株日蒸腾耗水量分别与日总太阳辐射和日均白天空气水气压亏缺进行指数饱和曲线拟合,拟合效果较好。采用2个方位和1个方位树干液流通量密度测算的整株蒸腾耗水量与采用4个方位的测算值分别相差约18%和30%,说明不同方位之间的液流差异可能成为蒸腾耗水测算的重要误差来源。