采用AFLP分析齐口裂腹鱼DNA指纹的技术探讨

AFLP（选择性扩增片断长度多态性）指纹技术被认为是当今最有效的DNA指纹分析技术之一,但是由于该技术操作流程较长,对模板DNA质量要求较高以及PCR反应成分等变数条件的影响,要获得高清晰的DNA指纹图谱比较困难。文中以齐口裂腹鱼为研究对象,对AFLP的分析条件作了一定的探索。结果表明：基因组DNA必须是高纯度、高分子量;酶切充分;PCR反应最佳条件为：引物E—XYZ,M—XYZ在体系中的浓度分别为1ng／mL,5ng／mL;dNTPs浓度为0．2mmol／L;MgCl2浓度为1．9mmol／L。银染时在NaOH条件下显色比在传统银染法中的Na2CO3条件下显色,图谱背景更干净,条带更清晰。     

马氏珠母贝选育系遗传变异的AFLP分析

利用AFLP技术对马氏珠母贝两个选育系DDS-G1和JCS-G1及大亚湾养殖群体(DDC)进行了遗传变异分析.5对选择性扩增引物共扩增谱带265条,DDC的多态位点比率为79.25%,DDS-G1为73.58%,JCS-G1为75.85%.两个选育系的单态位点明显增加,而Shannon's遗传多样性指数和Nei's基因多样性有所下降.三个组间的遗传距离为0.1615～0.2744,两个选育系间的遗传距离较大.研究结果表明,经过一代的选择,马氏珠母贝选育系的遗传结构和遗传多样性已发生改变,AFLP标记能够有效地监测选育系的遗传变异.     

Air quality biomonitoring: assessment of air pollution genotoxicity in the Province of Novara (North Italy) by using Trifolium repens L. and molecular markers

Mixed air pollutants are considered a major cause of DNA damage in living species. In this study Trifolium repens L. cv Regal was used as a bioindicator to assess the genotoxicity of air stressors in the Italian province of Novara. Two on-site biomonitoring experiments were performed during the spring and autumn of 2004. Test plants were exposed at 19 monitoring sites distributed homogeneously throughout the province, and each experiment lasted for a period of 6 weeks. Genotoxicity was evaluated with Amplified Fragment Length Polymorphism (AFLP) molecular markers. The results show the predominantly rural central-west region of the Novara Province to have the worst air quality with regard to genotoxicity. Analyses of geomorphology, land use and climatic factors suggest that the compromised air quality in the region could be attributed to wind strength and direction, transporting pollution from vehicular traffic on the A4 highway and from the urban/industrialized centres of Novara and Vercelli. Plant growth, changes in plant photochemical efficiency and the presence of ozone related leaf injuries were also measured to better interpret the results of genotoxicity. Statistical analyses show that although climatic factors such as light intensity and temperature influence plant growth, they do not contribute to atmospheric stressor-induced DNA damage. Further analyses indicated that, as expected, a mixture of genotoxic and non-genotoxic pollutants coexist in the Novara Province troposphere, and that the elevated ozone concentrations experienced during the study may have contributed to the DNA damage in the tested plants by enhancing genotoxicity via interaction with other air stressors.     

制作云南乳扇用酸乳清中乳杆菌的多样性分析

针对制作云南乳扇用酸乳清中的乳杆菌的来源及种属,对其多态性进行研究。采用AFLP技术和NTSYS-pc2.1软件对从云南大理白族地区牧民家庭制作乳扇用的的酸乳清中分离出的70株乳杆菌进行多样性分析。结果表明,AFLP扩增出的条带清晰,重复性好,多态性高。聚类分析表明,在0.89的水平上,70株乳杆菌共分为4个大群:高加索乳杆菌群Ⅰ、发酵乳杆菌群Ⅱ,瑞士乳杆菌群Ⅲ和植物乳杆菌群IV。其中,瑞士乳杆菌群Ⅲ中的乳杆菌占供试乳杆菌的65.7%,为优势菌群。进一步对优势菌群聚类,进行种内分型,以0.90为界,群Ⅲ被分为8个基因型。     

AFLP技术在玉米自交系类群分析研究中的应用

用ＡＦＬＰ分子标记技术对２３个应用较广泛的玉米自交系进行了遗传多样性分析。在ＡＦＬＰ分析当中,共采用了７个ＥｃｏＲＩ／ＭｓｅＩ引物组合,每个引物组合扩增出的条带数在５５－９６条之间。利用以上ＡＦＬＰ数据进行了聚类分析,将２３个玉米自交系分为３个群。结果表明,ＡＦＬＰ聚类图基本与系谱分析一致。ＡＦＬＰ技术是检测玉米自交系遗传多样性有效的分子标记技术。另外通过ＡＦＬＰ分析,找到了三个玉米自交系的特异Ａ     

The presence,genetic diversity and behaviour of Listeria monocytogenes in blue-veined cheese rinds during the shelf life

Blue-veined cheeses may allow Listeria monocytogenes survival and multiplication due to the biochemical characteristics of the cheese and the growth characteristics of the pathogen. Because of the availability of a considerable number of samples, we wanted this study to take a large view of the microbiota and of the incidence and genetic diversity of L. monocytogenes in blue-veined cheese rinds. Moreover, we wanted to determine if the pathogen present on the rinds at the end of ripening represented a risk to the consumer if the cheese should exceed the domestic storage limit imposed by Commission Regulation (EC) No 2073/2005 for food safety criteria.The rind microflora showed a high level of heterogeneity, and the incidence of L. monocytogenes (never found in the paste samples) was 55%, with an increased presence and concentration associated with a longer ripening time. Amplified Fragment Length Polymorphism (AFLP) was used to characterise L. monocytogenes isolated from blue-veined cheese rinds and highlighted the heterogeneity among the strains, demonstrating its suitability for studying the biodiversity of the pathogen in this environment. L. monocytogenes was able to grow during the shelf life of the cheese and was influenced by the refrigeration temperatures and the physicochemical characteristics of the cheese.     

AFLP技术在食品微生物研究中的应用

随着现代分子生物学技术的发展 ,对微生物学的研究也进入到分子、基因水平。扩增片段长度多态性(AFLP)是一种基于PCR的高分辨率DNA指纹技术 ,它能检测整个微生物基因组的多态性 ,并且结果可重复、分辨率高。近些年来 ,AFLP在细菌等微生物的遗传相关性鉴定、流行病学分型等上的应用得到迅速发展。文中介绍了AFLP技术的基本原理、反应流程 ,探讨了其在食品微生物研究领域中的应用现状和前景     

Cuticular Hydrocarbons and Aggression in the Termite Macrotermes Subhyalinus

Cuticular hydrocarbons are among the prime candidates for nestmate recognition in social insects. We analyzed the variation of cuticular hydrocarbons in the termite species M. subhyalinus in West Africa (Comoë National Park) on a small spatial scale (<1 km). We found considerable variation in the composition of cuticular hydrocarbons among colonies, with four distinct chemical phenotypes. Different phenotypes occurred within each of the four habitats. The difference between these phenotypes is primarily due to unsaturated compounds. A clear correlation between the difference of the hydrocarbon composition and the aggression between colonies was found. This correlation also holds in a multivariate analysis of genetic similarity (measured by AFLPs), morphometric distances (measured by Mahalanobis-distances), as well as geographic distances between colonies. In a more detailed analysis of the correlation between the composition of cuticular hydrocarbons and aggression, we found that no single compound is sufficient to explain variation in aggression between pairings of colonies. Thus, termites seem to use a bouquet of compounds. Multiple regression analysis suggested that many of these compounds are unsaturated hydrocarbons and, thus, may play a key role in colony recognition.     

Distribution of Penicillium commune isolates in cheese dairies mapped using secondary metabolite profiles, morphotypes, RAPD and AFLP fingerprinting

In an 8-year study of the diversity and distribution of Penicillium commune contaminants in two different cheese dairies, swab and air samples were taken from the production plants, the processing environment and contaminated cheeses. A total of 321 Penicillium commune isolates were characterized using morphotypes (colony morphology and colours) and secondary metabolite profiles. Based on production of secondary metabolites the P. commune isolates were classified into 6 groups. The genetic diversity of the P. commune isolates was assessed using randomly amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP). For a sub-set of 272 P. commune isolates RAPD analysis generated 33 RAPD groups whereas AFLP profiling revealed 55 AFLP groups. This study conclusively showed that the discriminatory power of AFLP was high compared to RAPD and that AFLP fingerprinting matched morphotyping. P. commune isolates with identical profiles using all four typing techniques were interpreted as closely related isolates with a common origin and the distribution of these isolates in the processing environment indicated possible contamination points in the cheese dairies. The coating process and unpacking of cheeses with growth of P. commune seemed to cause the contamination problems. Several identical P. commune isolates remained present in the processing environment for more than 7 years in both dairies.     

西藏野生油菜遗传多样性分析

利用10对AFLP引物及13对SSR引物,分析了43份来自西藏地区部分野生类型油菜种质的遗传多样性。10对AFLP引物共得到276条清晰的谱带,其中多态性带214条,多态性位点比率为77．5％,平均每对AFLP引物得到21．4条多态性带。13对SSR引物共扩增出57条带,其中多态性带51条,多态性位点比率为89．5％,说明西藏野生油菜遗传多样性丰富。通过对西藏野生类型油菜资源的遗传距离以及聚类分析,可将西藏野生类油菜分为两大类群,分别为白菜型和芥菜型野生油菜种质资源;两种分子标记适合揭示西藏野生油菜的遗传多样性。