葡萄逆境胁迫诱导启动子的克隆及表达分析

为了研究葡萄抗逆基因(CAN70200.1)的表达特性,采用PCR技术从葡萄中克隆了CAN70200.1基因上游一段长度为1 354 bp的启动子片段,命名为PCAN。采用Plant CARE和PLACE启动子在线预测工具分析表明,PCAN启动子序列具有CAAT-box、TATA-box基本的顺式作用元件和一些参与非生物胁迫、光和植物激素应答相关的顺式作用元件。为验证启动子的表达特性,将PCAN启动子连接到p CAMBIA1391Z载体GUS基因的上游,构建成植物表达载体p1391Z-CAN,并通过农杆菌介导法转化烟草,经PCR鉴定,获得转基因植株。对转基因烟草植株进行逆境胁迫处理发现,在干旱处理120 min后,PCAN启动子活性达到最强;而4℃低温处理30~60 min时,PCAN启动子活性达到最强,表明PCAN启动子具有低温和干旱胁迫诱导表达特性。     

葡萄酒中总铁和Fe2+的改良菲洛嗪法快速测定

通过优化菲洛嗪法测定葡萄酒中总铁的检测条件,以及探讨Fe~(3+)共存时菲洛嗪用量对Fe~(2+)测定的影响,建立一种简易可靠、能够直接测定葡萄酒中总铁和Fe~(2+)的方法。结果显示:改良菲洛嗪法测定葡萄酒中的总铁,在0.25~2.00 mg/L质量浓度范围内线性关系良好,检测限为0.011 5 mg/L,加标回收率为94.31%~104.34%,变异系数范围为0.95%~2.43%;然而,共存的Fe~(3+)会影响对葡萄酒中Fe~(2+)的测定,适合的显色剂用量是菲洛嗪与总铁摩尔比为7,在此条件下,Fe~(2+)质量浓度的拟合准确度为101.98%~113.50%。利用该法测定葡萄酒样品的结果表明,葡萄酒中以Fe~(2+)为主,其在红葡萄酒中的百分比相对较高。总之,改进的方法简单易行、准确可靠,能够满足葡萄酒中铁价态的快速定量测定,适于在葡萄酒生产中推广应用。     

0~#柴油和东海平湖原油对黑鲷血细胞DNA损伤的研究

采用单细胞凝胶电泳技术(SCGE),以砂滤过的清洁海水为对照组,研究0~#柴油(0.015 mg·L~(-1)、0.03 mg·L~(-1)、0.06mg·L~(-1))和东海平湖原油(0.03 mg·L~(-1)、0.06 mg·L~(-1)、0.12 mg·L~(-1))在不同暴露时间(3 d、7 d、15 d和21 d)对黑鲷血细胞核DNA损伤作用。结果显示,黑鲷血细胞DNA损伤程度与两种石油有显著的剂量-效应和时间-效应关系。实验第3天,0~#柴油和东海平湖原油试验组就检测到DNA损伤,主要以1级损伤为主。随0~#柴油和平湖原油浓度的增高和胁迫时间的延长,彗尾DNA相对含量(Tail DNA%)和彗尾长度/核直径(TL/D)值均呈上升趋势,DNA损伤级别逐渐升高。恢复实验结束后,中、高浓度组0~#柴油(0.03 mg·L~(-1)、0.06 mg·L~(-1))和平湖原油(0.06 mg·L~(-1)、0.12 mg·L~(-1))的彗尾DNA相对含量(Tail DNA%)和彗尾长度/核直径(TL/D)值无法恢复至对照组水平(P0.05)。从DNA损伤程度来推测,0~#柴油的生物毒性大于平湖原油。研究表明,鱼类血细胞DNA损伤可作为监测海洋石油污染的生物标志物。     

SRBSDV结构蛋白P10和非结构蛋白P9-1抗体的制备及应用

为丰富南方水稻黑条矮缩病毒(Southern rice black-streaked dwarf virus,SRBSDV)的预测预报体系,本研究利用Gateway原核表达系统,将SRBSDV结构蛋白P10和非结构蛋白P9-1进行了大量表达,纯化的表达蛋白分别免疫新西兰大白兔(Oryctolagus cuniculus)获得抗血清,抗血清的间接酶联免疫吸附分析(indirect enzyme-linked immunosorbent assay,in-ELISA)效价分别为1∶6 400和1∶3 200,Western blot检测结果表明,制备的2种抗体均具特异性.利用免疫捕获RT-PCR(immunocapture RT-PCR,IC-RT-PCR)和斑点酶联免疫吸附分析(dot enzyme-linked immunosorbent assay,dot-ELISA)方法比较两种抗体检测水稻(Oryza sativa)和白背飞虱(Sogatella furcifera,WBPH)带毒率的应用效果,结果表现一致,说明SRBSDV结构蛋白P10和非结构蛋白P9-1均可用于病毒的田间检测,为病害的预测预报奠定了基础.     

Investigation into the use of the amylase SD assay of milled wheat extracts as a predictor of baked bread quality

α-Amylase content in milled wheat is a key quality parameter in the baking industry. This metric is usually determined using the Hagberg Falling Number (FN) method, the results of which can be used to segment grains into a range of quality categories. The authors describe herein an alternative to the traditional FN assay, namely the Amylase SD assay, as a predictor of baked bread quality. 144 Wheat grain samples with known FN values were analysed using the Amylase SD assay and a simple model for the conversion of Amylase SD units into “predicted FN” (PFN) values has been described. To test the usefulness of this approach, 11 separate wheat grain samples were analysed using the FN method, the Amylase SD assay, and also for gluten content. These grain samples were then milled to produce flour which was re-analysed using the Amylase SD assay and then formulated into dough that was subjected to Mixolab analysis, before being baked to produce pan bread. Correlations of baked bread volume and common Mixolab parameters to both FN and PFN values have been investigated for each sample. The Amylase SD assay has been shown to be a potentially promising alternative to the FN method for use as a baked bread quality predictor.     

Development of a real-time fluorescence loop-mediated isothermal amplification assay for rapid and quantitative detection of Fusarium mangiferae associated with mango malformation

Fusarium mangiferae is a major causal agent of mango malformation disease (MMD) worldwide. Rapid and accurate detection of the causal pathogen is the cornerstone of integrated disease management. In this paper, a real-time fluorescence loop-mediated isothermal amplification assay (RealAmp) was developed for quantitative detection of F. mangiferae in China. The LAMP primer set was designed based on a RAPD marker sequence and positive products were amplified only from F. mangiferae isolates, but not from any other species tested, showing a high specificity of the primer sets. The detection limit was approximately 2.26 × 10⁻⁴ ng/μl plasmid DNA when mixed with extracted mango DNA. Quantification of the pathogen DNA of MMD in naturally collected samples was no significant difference compared to classic real-time PCR Additionally, RealAmp assay was visual with an improved closed-tube visual detection system making the assay more convenient in diagnostics.     

永停滴定法测定磺胺间甲氧嘧啶钠注射液含量的不确定度评定

以磺胺间甲氧嘧啶钠注射液为例,阐述永停滴定法含量测定结果的不确定度评定。通过建立数学模型,分析不确定度来源,对各个不确定度分量进行了评估,计算了扩展不确定度,含量测定结果可表述为(95.3±0.8)%(k=2)。结果表明,重复性测量、校准和温度差异引入的不确定度为测量结果不确定度的主要来源。     

The ultra-sonication of minerals in swine feed

A sample preparation method based on ultrasound assisted-extraction (UAE) of Ca, Mg and P from swine feed has been described. The experiment was performed to cover the variables influencing the sonication process and, the method validation using standard reference material. Final solutions obtained upon sonication were analyzed by flame atomic absorption spectrometry (for Ca and Mg) and by UV–vis spectrophotometry (for P). The best conditions for metal extraction were as follows: sample mass: 100 mg in 20 mL 0.10 mol/L HCl, a particle size: <60 μm, sonication time: 5 cycles of 10 s and ultrasound power: 102 W. The UAE method was applied in digestibility assays in different piglet feeds and their results showed that it is highly comparable (P > 0.05) to the other methods used for such purposes, as block digestion, and offered a Ca, Mg and P method of quantification limit of 10.6, 12.4 and 14 mg/kg, respectively. The major advantages of the UAE method compared to other methods are the high treatment rate, low reagent usage in the extracts and, it does not generate toxic residues that might negatively affect human health and the environment, accompanied by good precision and accuracy.