蜡梅花瓣基因组DNA提取及RAPD-PCR反应体系优化

以蜡梅花瓣为试材,用改良CTAB法提取基因组DNA,并设置不同浓度梯度对每个PCR反应因子进行相应试验,结果表明,改良CTAB法提取蜡梅花瓣基因组纯度高、质量好。并采用正交试验设计的方法,对蜡梅RAPD-PCR条件进行了优化,结果表明,最佳的蜡梅RAPD-PCR反应体系(20μL)为:1×buffer,1.0 U TaqDNA聚合酶,2.0 mmol/L MgCl2,0.15 mmol/L dNTPs,0.5μmol/L引物和模板DNA 30～40 ng;适宜的扩增条件为:94℃预变性3 min;94℃变性30 s,38℃退火30 s,72℃延伸90 s,38个循环;72℃延伸7 min,4℃保存。     

9种鹤类DNA条形码的研究

为了研究9种鹤类的DNA条形码,采用DNA条形码鸟类通用引物扩增中国4种鹤类（22个个体）的线粒体细胞色素C氧化酶亚基I（COI）基因。从GenBank及BOLD Systems下载13条鹤类的COI基因序列,对35条COI序列进行分析,构建NJ树和MP树,探讨DNA条形码对鹤类的识别和鉴定。结果显示,丹顶鹤群体中,除D1和B18号个体外,均被归为同一类群;白鹤群体中,编号为AY567881的个体与黑鹤划为同一类群;其余鹤类的系统发育分析结果均与形态学分类结果相同。由于丹顶鹤样本均取自沈阳森林野生动物园,所以是同一物种或是近缘物种杂交的可能性最大;白鹤较大的种内差异则可能是地理位置或是其他原因。     

基于atpF-atpH序列鉴定稻属植物

为从分子水平上鉴定稻属,对稻属9种32份植物材料DNA条形码基因atpF-atpH进行PCR扩增并对PCR扩增产物测序,以近缘种植物为外类群建立系统进化树及序列差异分析,设计了鉴定稻属的特异性引物。结果表明,应用该特异性引物对稻属材料进行扩增,均获得与预期大小一致的113bp特异性目的片段,而对非稻属材料的扩增结果均呈阴性。系统进化树也显示出atpF-atpH基因可以很好将稻属区分开。这说明在条码基因差异位点分析的基础上,设计特异分子标记可用于稻属的鉴定,为珍贵物种的口岸鉴定提供了检测方法。     

Relation between pathogenicity and genetic variation within <Emphasis Type="Italic">Plasmodiophora brassicae</Emphasis>

The relation between diversity of pathogenicity on clubroot-resistant (CR) cultivars of Chinese cabbage (Brassica rapa subsp. pekinensis) bred in Japan and DNA polymorphisms in 17 populations of Plasmodiophora brassicae from cruciferous plants was examined by inoculation tests and random amplified polymorphic DNA (RAPD) analysis using 18 arbitrary primers. Four pathotypes (A–D) were identified after inoculation of six CR cultivars of Chinese cabbage in the 17 populations from cruciferous crops. A relatively high level of genetic diversity was also detected among these populations in the RAPD analysis. Although the four pathotypes could not be clearly differentiated using the RAPD data, most populations of three pathotypes had a consistent location on the dendrogram. All pathotype B (virulent on five cultivars except Utage 70) and D (avirulent on all cultivars) populations, which were common in incompatible interactions with cv. Utage 70, were located in a single subcluster. All five pathotype C populations (virulent only on cv. Utage 70) except for one population grouped in another single subcluster. Because four pathotype A populations (virulent on all six cultivars, races 4 and 9) fell in different subclusters, the populations may be genetically polyphyletic. Populations from cruciferous weed Cardamine flexuosa differed remarkably from those from cruciferous crops in pathogenicity on common cultivars of Chinese cabbage and turnip and C. flexuosa, but they grouped in a single cluster with all race 9 populations from crops. Race 9 populations from crops may thus be closely related to populations from the weed rather than to races 1 and 4 from crops.     

Within-field variability of Fusarium head blight pathogens and their associated mycotoxins

Within-field variability in the Fusarium head blight (FHB) and its associated mycotoxins was studied in four European countries. At each of 14 sites, each FHB pathogen and associated mycotoxins were quantified in 16 quadrat samples at harvest. Overall, the incidence of quadrat samples with detectable and quantifiable pathogen DNA was significantly lower in the grain than in the corresponding chaff. Deoxynivalenol (DON) was the most frequently detected toxin in the samples and its accumulation was most strongly associated with the presence of Fusarium graminearum. Nivalenol (NIV) accumulation was significantly associated only with the presence of F. culmorum. Zearalenone (ZON) accumulation was strongly associated with the presence of all three pathogens (F. graminearum, F. culmorum and F. poae). The levels of both DON and ZON concentrations were positively related to the amount of F. graminearum DNA in the grain or in the chaff. The presence/absence of FHB pathogens within a single quadrat appeared to be independent of each other. The presence of a particular FHB pathogen and the amount of its DNA, as well as the associated mycotoxin(s), varied greatly among samples at each site. This study demonstrated the large extent of within-field variability of FHB and its associated mycotoxins, and the importance of representative sampling in FHB studies.     

Lead (Pb) accumulation,oxidative stress and DNA damage induced by dietary Pb in tilapia (Oreochromis niloticus)

With the increasing occurrence of dietary lead (Pb) contamination in aquaculture, a better understanding of the toxic effects of dietary Pb on aquatic animals is needed. Tilapia (Oreochromis niloticus) were exposed to dietary Pb at concentrations of 0, 100, 400 and 800 μg g^?1 dry weight for 60 days, and Pb accumulation in tissues and blood, oxidative stress in posterior kidney and DNA damage in peripheral blood cells were investigated. The results showed that dietary Pb exposure resulted in significant Pb accumulation in tissues and blood, which increased with the dietary Pb concentrations. Pb accumulated in sampled tissues in the following order: posterior kidney>bone>liver>gill>spleen>testis>muscle>brain. Dietary Pb caused a significant increase in the malondialdehyde level and a significant decrease in the total antioxidant capacity content when compared with the control group (P<0.05), accompanied by concentration‐dependent decreases in the glutathione content, glutathione peroxidase and superoxide dismutase activities. Pb dose‐dependent DNA damage was observed in peripheral blood cells of tilapia exposed to dietary Pb. The results suggest that dietary Pb exposure can induce significant Pb accumulation in tissues and blood, followed by oxidative stress in posterior kidney and DNA damage in peripheral blood cells of tilapia.     

Seafood mislabelling: comparisons of two western European case studies assist in defining influencing factors,mechanisms and motives

The global seafood industry, influenced by consumer demand, is closely linked to the global fishing industry, which determines the variety of fish available for consumption. The recently revealed issue of seafood mislabelling threatens to weaken this link by removing consumer power to influence patterns of fisheries exploitation through informed choice. Recognizing this, there is an urgent need to go beyond the mere documentation of the phenomenon and learn more about the origins of this problem and the nature of factors influencing its occurrence to develop solutions. In an attempt to understand seafood mislabelling more thoroughly in Europe, 226 cod products were purchased from Ireland and the UK, genetically identified using a DNA barcoding technique (COI barcoding gene), and species identification results were compared against product labels. Cod mislabelling proved more severe in Ireland than in the UK (28.4% vs. 7.4%). Moreover, whereas data show that in Ireland, cheaper species are sold as cod, in the UK, threatened Atlantic cod (Gadus morhua) may be sold as ‘sustainably sourced’ Pacific cod. Considering these countries operate under the same EU policies for seafood traceability and labelling, it is likely that this situation has been influenced by heightened consumer awareness in the UK, which has created an environment where mislabelling is discouraged. In addition to identifying samples, traceability information from packaged cod was used to trace products back to supplying companies. Although inconclusive in determining blame, this exercise has demonstrated that using traceability information can add explanatory power when attempting to determine responsibility for the occurrence of mislabelling.     

改良CTAB法在核桃叶片基因组DNA提取中的应用研究

核桃叶片中酚类、多糖和蛋白含量丰富,影响了基因组DNA的提取。以CTAB法为基础进行了改良,并对核桃叶片基因组DNA进行提取。结果表明,通过在研磨时添加PVPP后的改良CTAB法不仅操作简单,经济高效,而且去除多糖和多酚类物质彻底,能成功用于核桃的基因克隆研究,在核桃DNA提取中值得大力推广使用。     

DNA提取方法对转基因甘蔗PCR检测的影响

研究了不同提取方法和不同生长时期甘蔗提取的DNA对转基因甘蔗植株PCR检测的影响。结果表明,幼苗时期提取的甘蔗叶片模板DNA质量高于田间成熟植株;甘蔗幼苗时期的PCR检测易于成熟植株;CTAB法、SDS法和CTAB-SDS法提取的甘蔗叶片DNA均能满足幼苗期甘蔗抗性植株的PCR检测要求;对成熟转基因甘蔗植株而言,以CT...     

4种滑叶铁线莲基因组DNA提取方法比较(英文)

[目的]对4种滑叶铁线莲基因组DNA提取方法进行比较研究,建立滑叶铁线莲最适的DNA提取方法。[方法]以滑叶铁线莲叶片为材料,比较改良CTAB法Ⅰ、改良CTAB法Ⅱ、改良CTAB法Ⅲ、改良SDS法这4种基因组DNA提取法在提取的DNA纯度、浓度和提取时间等方面的不同。[结果]4种方法都可提取滑叶铁线莲基因组DNA。改良CTAB法Ⅰ提取DNA纯度最高,但浓度最低且提取时间最长;改良SDS法提取DNA浓度最高,所需时间较短,但纯度较低;改良CTAB法Ⅲ提取所需时间最短。[结论]建立了铁线莲最适DNA提取方法,为运用分子生物学手段对其研究提供支持。