大白菜杂交种中熟5号纯度的RAPD鉴定

利用RAPD-PCR分子标记技术,对大白菜品种中熟5号及其亲本的DNA指纹进行了分析研究,从200个随机引物中筛选出互补带明显的引物S33,构建了相应的DNA指纹图谱.通过对100株中熟5号进行纯度鉴定,结果只有2个有母本特异带,与大田检测的结果完全一致,杂交率达98%.     

荷斯坦牛耐热性RAPD和SCAR标记的研究

以52头高产耐热组和41头高产不耐热组中国荷斯坦牛的DNA为材料,选用200条RAPD引物,利用RAPD-PCR技术,对荷斯坦牛的耐热性进行研究。结果发现,使用S441引物和S463引物在高产耐热组中分别发现了581 bp（RAPD-S441标记）和680 bp（RAPD-S463标记）2条特异性片段。通过测序和序列比较显示：581 bp的片段与KCNN2基因的同源性达90%;680 bp片段与NW_001024067.1｜BtUn_WGA9442_2的同源性达99%。根据已知KCNN2基因的功能,初步判断KCNN2基因为影响耐热性能的候选基因。得到GenBank登录号EF123743和EF123744。利用NCBI的ORF finder在581 bp中没有发现ORF,与预测其是KCNN2基因内含子3的结论相符合。在680 bp中发现4个开放阅读框（ORF）,分别为ORF1、ORF2、ORF3、ORF4。这4个阅读框分别编码48AA、34AA、45AA、34AA。根据RAPD-S441、RAPD-S463标记的测序结果,利用Premier 5.0软件设计对应的SCAR引物。成功将RAPD-S441标记转化为耐热SCAR标记。     

Different methods for the evaluation of species of the genus Mesaphorura (Collembola, Tullbergiinae)

Field studies and laboratory investigations based upon cultures, isoenzyme electrophoresis and RAPD-PCR were carried out to get a better understanding of the value of morphospecies within the genus Mesaphorura.Culturing of seven Mesaphorura species under constant as well as variable conditions provided a basis for evaluation of morphological characters, but did not yield any statistically supportable evidence that intraspecific variability reached interspecific delimitations.In enzyme tests, esterases gave the best results. The search for species-specific band patterns is, however, time and material consuming. DNA investigations using RAPD-PCR resulted in band patterns that appeared to be species specific. Nevertheless, these require further investigations.None of the studies carried out gave any conclusive evidence for invalidation of the morphologically defined species of the genus Mesaphorura.     

Reliable detection of the fungal pathogenfusarium oxysporum f.sp.albedinis, causal agent of bayoud disease of date palm, using molecular techniques

Bayoud, caused by the soilborne fungusFusarium oxysporum f.sp.albedinis (FOA), is the most serious disease of date palm. Since the disease is located in the North African countries of Morocco and Algeria, and advancing steadily eastwards, the ultimate goal is to prevent spread of the pathogen to other date-growing areas in the region and farther afield. Molecular diagnostic techniques have been developed for detection of FOA. In view of the fact that the fungus does not exist in Israel, DNA of FOA was obtained to determine the reliability of these methods for diagnostic purposes. Random amplified polymorphic DNA was not reliable enough for differentiation between FOA and various pathogenic and saprophyticFusarium isolates. However, the polymerase chain reaction utilizing FOA-specific primers was accurate and enabled amplification of a unique band specific to FOA DNA alone, and not that of the other tested pathogenic and saprophyticFusaria. The availability of a rapid and reliable diagnostic tool for detection of FOA will enable the Plant Protection and Inspection Services of the Israel Ministry of Agriculture to test date palm tissue for the presence of the pathogen. Contribution no. 513/00 from the Inst. of Plant Protection, ARO, The Volcani Center, Bet Dagan, Israel.     

蒌蒿DNA提取、RAPD优化及引物筛选初报

为了能从分子水平探讨蒌蒿资源的遗传多样性、蒿属的系统分类和种质资源的鉴定,在此,报道采用小量法（SDS法）和大量法（CTAB法）提取蒌蒿DNA,建立优化的RAPD-PCR体系,并进行引物初步筛选。结果显示,两种抽提方法都可有效抽提出高质量的DNA,大量法得率要明显高于小量法,但小量法抽提的DNA也足够RAPD-PCR反应几十至上百次,不同的实验室可根据实验的需要进行选择。对两种方法抽提的DNA,都进行PCR扩增体系梯度摸索,最优的RAPD-PCR反应条件为：25μl反应体系中,含DNA模板约20ng,10×TaqDNA聚合酶缓冲液2.5μl,2μldNTPs（2.5mM）,Mg2＋2μl（25mM）,TaqDNA聚合酶0.15μl（5U/μl）,引物0.5μl（25μM）,其余以双蒸水补充。在剔除了重复性差,条带模糊和单态的引物后,有九个引物表现稳定,扩增出来的条带清晰、多态性高。     

苹果基因组DNA的快速提取

提出了一种适用于苹果（Malus pumila Mill)RAPD分析的基因组DNA的快速提取方法。用本法提取的DNA质量高，简便，快速，经济，每天可提取200个DNA样品，每个DNA样品可供50－100次PCR反应，用10个碱基随机引物进行PCR扩增，95％以上的样品都才扩增出3－10条清晰的泳带。     

Genetic differentiation of the parthenogenetic soil collembolan Isotoma notabilis along a copper gradient based on random amplified polymorphic DNA

Five species of collembolans were collected at 24 sampling areas in a copper polluted field. The most abundant species was Isotoma notabilis found at half the sampling sites, in total 490 individuals. Two hundred and fifty-two specimens of these, representing seven sampling areas were analysed by means of random amplified polymorphic DNA using three decamer primers. Two primers each revealed eight phenotypes and the third showed six phenotypes. Distribution of the phenotypes was not homogenous within the sampling area. This result could not be explained as an outcome of either the copper content in the soil or the grass biomass, but was more likely an effect of colonisation from the areas surrounding the field.     

中华蜜蜂王浆中DNA的提取方法研究

在国内外首次以中华蜜蜂新鲜王浆为材料,采用改进的苯酚-氯仿法,提取出其DNA并对其含量及纯度进行测定。结果表明:中华蜜蜂新鲜王浆中DNA的含量为66.29~105.34μg/g,平均含量为(82.95±10.39)μg/g,平均纯度(A260/A280)为1.69±0.08,以其作为模板进行RAPD-PCR扩增,扩增的条带较清晰、明亮、稳定性好。     

An assessment of the genetic diversity within a collection of<Emphasis Type="Italic">Saccharum spontaneum</Emphasis> L. with RAPD-PCR

A local collection of 33Saccharum spontaneum L. clones and two sugarcane cultivars (LCP 82-89 and LCP 85-384) were assessed for genetic variability using random amplified polymorphic DNA (RAPD)-PCR. A total of 157 polymorphic RAPD-PCR bands were scored with 17 primers. The number of RAPD-PCR products per primer ranged from four to 16. The data were analyzed with two multivariate analysis software programs, NTSYSpc and DNAMAN®. Although these two programs yielded similar results, a bootstrapped phylogenetic tree could only be generated with the DNAMAN® software. A substantial degree of genetic diversity was found within the localS. spontaneum collection. Pairwise genetic homology coefficients ranged from 65% (SES, 196/Tainan 2n = 96) to 88.5% (IND 81-80/IND 81-144). LCP 82-89 and LCP 85-384 shared a greater similarity (82%) than either was to any clone ofS. spontaneum (ranging from 60.5 to 75.2%). The 33S. spontaneum clones were assigned to eight groups independent of their geographic origin or morphology, while the two sugarcane cultivars were assigned to the ninth group. All but two pairs ofS. spontaneum clones could be distinguished by a single RAPD primer OPBB-02. The use of a second primer, either OPBE-04 or Primer 262, separated allS. spontaneum clones. One amplification product from the RAPD primer OPA-11, OPA-11-336, proved to be cultivar-specific and has been adopted for use in our breeding program. Information from this study would help conserve the genetic diversity ofS. spontaneum. Disclaimer: Product names and trademarks are mentioned to report on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by USDA does not imply the approval of the product to the exclusion of others that may also be suitable. The experiments reported comply with the current laws of the USA.     

松口蘑菌丝分离及其RAPD—PCR鉴定

采用组织分离法,对东宁县的松口蘑进行了分离培养。通过显微观察及RAPD—PCR技术对松口蘑子实体及其相应的培养物之间进行了亲缘关系鉴定。结果表明,菌褶为松口蘑组织分离的最适宜部位,其最佳分离培养基为冈叮,即PDA＋黑木耳二级种浸出液＋麦麸。RAPD—PCR结果显示,松口蘑子实体与分离物之间的DNA指纹相似系数在0．97～1．00之间,表明松口蘑子实体与其相应的培养物在种质上是一致的。