棉花枯萎病菌异核体不同核型分离子表观特性及其核DNA的RAPD分析

切割野生型棉花枯萎病菌Fusariumoxysporumf.sp.vasinfectum菌落的菌丝尖端,得到菌株Ag149,该菌株是一个异核体,其菌落出现明显角变,自角变处经单孢分离和继代培养,得到三种表观性状稳定的分离子Ag149-I、Ag149-II和Ag149-III。它们在色素产生、菌落质地、孢子产量以及致病性上存在明显差异,但对这三种核型分离子核DNA进行RAPD分析,未发现明显差异。     

棉花枯萎病菌异核体不同核型分离子表观特性及其核DNA的RAPD分析

切割野生型棉花枯萎病菌Ｆｕｓａｒｉｕｍ ｏｘｙｓｐｉｒｕｍ ｆ．ｓｐ．ｖａｓｉｎｆｅｃｔｕｍ菌落的菌丝尖端,得到菌株Ａｇ１４９,该菌株是一个异核体,其菌落出现明显角变,自角变处经单孢分离和继代培养,得到三种表观性状稳定的分离了：Ａｇ１４９－Ⅰ、Ａｇ１４９－Ⅱ和Ａｇ１４９－Ⅲ。它们在色素产生、菌落质地、孢子产量以及致病性上存在明显差异,但对这三种核型分离子核ＤＮＡ进行ＲＡＰＤ分析,未发现明显差异。     

棉花枯萎病菌异核体的三个不同核型分离子DNA碱基组成及18S rDNA序列

从棉花枯萎病菌（Fusarium oxysporum f.sp.vasinfectum）异核体菌株Ag149的菌落我变处分离得到3个培养特征和致病性差异明显的单孢分离子。分别测定了这3个分离子的总DNA和核DNA的碱基组成,以及它们的18S rDNA的部分序列（1477个碱基对）。它们的总DNA（G＋C）mol%差异在0.1%-0.4%之间;核DNA（G＋C）mol%的差异在0.4%-0.8%范围内;而18S rDNA的部分序列则完全相同。     

棉花枯萎病菌异核体的三个不同核型分离子DNA碱基组成及18S rDNA序列*

从棉花枯萎病菌 (Fusariumoxysporum f.sp .vasinfectum)异核体菌株Ag1 49的菌落角变处分离得到 3个培养特征和致病性差异明显的单孢分离子。分别测定了这 3个分离子的总DNA和核DNA的碱基组成 ,以及它们的 1 8SrDNA的部分序列 ( 1 477个碱基对 )。它们的总DNA (G +C)mol%差异在 0.1 %～ 0.4%之间 ;核DNA (G +C)mol%的差异在0.4%～ 0.8%范围内 ;而 1 8SrDNA的部分序列则完全相     

棉枯萎病菌异核体及其不同表型分离子在基因转录水平上的差异

采用银染mRNA差异显示技术,研究了棉花枯萎病菌异核体菌株(Ag149)及其两个不同表型分离子(Ag149-Ⅰ和Ag149-Ⅲ)之间差异表达的基因,观察到在300～700bp之间出现了19个差异条带,经反向RNA印迹,证实其中2条差异带为阳性：编号为C6的差异条带在Ag149和Ag149-Ⅰ菌株中呈高表达,而编号为G5的差异条带在菌株Ag149和Ag149-Ⅲ中呈高表达。这两条差异片段经测序后在GenBank中分别进行同源比较,发现由C6片段编码的氨基酸序列与多种生物(包括动物、植物和微生物)NADH脱氢酶的第6个亚基有不同程度的同源性(30％～70％);而编码G5片段的氨基酸序列与龟裂链霉菌(Streptomyces rimosus)的OtrB(tetracycline efflux protein)蛋白有35％的同源性。首次证明在异核体菌株及其不同表型分离子之间存在基因转录水平差异,为深入研究丝状真菌异核体形成的分子遗传机制提供了线索。     

球孢白僵菌单孢子分离株在继代培养过程中菌落局变的遗传分析

研究了球孢白僵菌Beauveria bassiana单孢株继代过程中的菌落局变现象,并通过AFLP分析了角变子与原菌株间在分子水平上的差异。结果表明,野生型出发菌株Bb13的单孢分离株13S5和13S8在继代培养过程中均发生生长速率加快和产孢量下降现象。13S5在前5代,13S8在前6代产孢量有所下降但不显著,至第10代时产孢量比第1代分别下降了81.7%和69.0%。菌株角变现象在继代培养5-6代后表现明显,而13S8角变子出现的频率更高。AFLP指纹图谱分析表明,用20个引物组扩增出的98个位点中,两个角变子与野生型菌株间有12条差异条带,变异率达12.2%。由此证明单孢子分离株在继代培养中发生菌落局变后遗传物质已产生了变异。     

球孢白僵菌继代培养中的菌落局变现象

通过球孢白僵菌两个野生型菌株及其单孢株在不同培养基、温度、湿度和光照条件下继代培养,结果产生3种不同类型的菌落局变.局变产生的分离子多表现为菌落瘩薄或气生菌丝陡长、产孢量下降、生长速率增加等现象.培养基、温度、湿度和光照等因素对菌落局变有明显诱导作用.湿度为15℃,相对湿度较低及全光照条件下于SDAY培养基上培养,局变发生频率最低.在实验室继代培养中,野生型菌株在4-14代内即可发生局变而被局变分离子所取代.引人注意的是,单孢株局变产生的分离子同时伴随着棕黄色或红棕色素的分泌,仅以菌落形态和颜色等培养特征作为球孢白僵菌种的划分依据是不可靠的.文中提出了菌落局变发生的遗传变异可能机制.     

蛹虫草表型多态性对子实体产生及虫草菌素的影响

为选育高产子实体和虫草菌素的蛹虫草菌株,从野生蛹虫草中分离单孢子并进行分型,之后进行产子实体实验;同时对蛹虫草子实体进行了产孢结构的观察,并用高效液相色谱仪检测了子实体和培养基中的虫草菌素。结果表明,菌落背面颜色为橙铬色的菌株容易产生子实体;出发(原代)菌株所产子实体在形态上更接近野生蛹虫草,而角变株的子实体畸形;出发菌株产子实体能力要比该菌株未发生角变部分分离株和角变部分分离株都强。在虫草菌素的产量上也以出发菌株为高,表明表型多态性对蛹虫草产子实体和虫草菌素有很大的影响。这对于优势菌种的筛选和生产实践有借鉴意义。     

用秋水仙碱诱发的金霉菌异型细胞质基因突变型研究

构巢毒菌(Aspergillu nidulans)的异型细胞质基因突变,具有在无性繁殖中表现持久的分离现象、不能获得纯系的突变型菌株以及异核体分离子的异型细胞质基因的表型基本上不受核的影响等的遗传特征。金霉菌(Strcptomyces aureofaciens)经秋水仙碱处理后,也发现类似构巢霉菌的异型细胞质基因突变现象。每一个突变型的菌落在生长过程都发生形态上的分离现象,能产生两个或两个以上的正常型角变。突变型菌落中央部分的分生孢子具有持久性的形态上的分离现象,即它所形成的菌落永远是突变型的菌落。但角变部分的分生孢子所形成的菌落都是正常型菌落。     

一种简捷的分离杜鹃花类菌根真菌的方法

目的：简化杜鹃花类菌根真菌的分离方法,快速得到菌株。方法：选择并改良马丁氏-孟加拉红培养基,调整其pH值约为5．0,0．3～0．5cm根段平铺于平板中培养分离菌株,培养条件为25℃黑暗培养12～15d。结果：得到了菌落特征有明显差异的9种类型138个菌根真菌菌株,其中7类菌株在人工条件下回接云锦杜鹃形成了菌根结构。结论：实验证明改良的分离方法操作简单快捷,是杜鹃花类菌根真菌分离的一种可靠方法。     

网纹瓜单细胞固-液双层培养及体细胞无性系建立(简报)

网纹瓜(Cucumis melo Var.reticulatus)是名贵的优质甜瓜,为葫芦科、甜瓜属植物。我国引种后生长良好,产量高,经济效益好,每年从日本购进大量杂交种子。但由于种子不纯,基因型差异较大,植株不整齐,产量不稳定,而且价格昂贵。因此,选育适合我国种植的     

FRACTIONATION OF NUCLEAR,CHLOROPLAST, AND MITOCHONDRIAL DNA FROM POLYSIPHONIA BOLDII (RHODOPHYTA) USING A RAPID AND SIMPLE METHOD FOR THE SIMULTANEOUS ISOLATION OF RNA AND DNA1

Extraction of nucleic acids from red algae is complicated by the presence of phycocolloids. For this reason, methods used for nucleic acid isolation from other organisms are not always amenable to use with red algal preparations; modifications in some cases lead to protocols that are time consuming and complicated, often requiring large amounts of algal tissue for starting material. Here we describe the isolation of both RNA and DNA followed by fractionation and identification of nuclear, chloroplast, and mitochondrial DNAs from a single preparation of Polysiphonia boldii Wynne and Edwards using a simple method that yielded approximately 100 μg of total RNA and 20 μg of total DNA from 1 g of frozen powdered algae. The potent protein denaturant guanidinium thiocyanate and the detergent sarkosyl were used to gently lyse the cells and organelles and immediately inhibit nuclease activity in the extract. The nucleic acids were isolated by ultracentrifugation into a dense solution of CsCl; the RNA was recovered as a pellet and the DNA as a band within the CsCl solution. Agarose gel electrophoresis of the total RNA showed discrete ribosomal RNA bands, indicating little nonspecific degradation. The nuclear, chloroplast, and mitochondrial DNAs were fractionated by density gradient ultracentrifugation in the presence of the DNA binding dye, bisbenzimide H (Hoechst 33258), which binds preferentially to DNA with a high A + T:G + C ratio, thus altering its density to a greater degree than it does that of DNA with a lower nucleotide ratio. The three fractions were identified by Southern blot analysis using heterologous gene probes specific for the different genomes. The protocol should be applicable to different types of algae. The simple nucleic acid isolation step can be performed on multiple samples simultaneously without subsequent fractionation of DNA, allowing comparison of DNA from different individuals, populations, or species.     

High nuclear genetic diversity, high levels of outcrossing and low differentiation among remnant populations of Quercus petraea at the margin of its range in Ireland

• Background and Aims Quercus petraea colonized Irelandafter the last glaciation from refugia on mainland Europe. Deforestation,however, beginning in Neolithic times, has resulted in small,scattered forest fragments, now covering less than 12 000 ha. • Methods Plastid (three fragments) and microsatellitevariation (13 loci) were characterized in seven Irish populationssampled along a north–south gradient. Using Bayesian approachesand Wright’s F-statistics, the effects of colonizationand fragmentation on the genetic structure and mating patternsof extant oak populations were investigated. • Key Results All populations possessed cytotypes commonto the Iberian Peninsula. Despite the distance from the refugialcore and the extensive deforestation in Ireland, nuclear geneticvariation was high and comparable to mainland Europe. Low populationdifferentiation was observed within Ireland and populationsshowed no evidence for isolation by distance. As expected ofa marker with an effective population size of one-quarter relativeto the nuclear genome, plastid variation indicated higher differentiation.Individual inbreeding coefficients indicated high levels ofoutcrossing. • Conclusions Consistent with a large effective populationsize in the historical migrant gene pool and/or with high geneflow among populations, high within-population diversity andlow population differentiation was observed within Ireland.It is proposed that native Q. petraea populations in Irelandshare a common phylogeographic history and that the presentgenetic structure does not reflect founder effects.     

A comparison of two pre-enrichment media prior to immunomagnetic separation for the isolation of E. coli O157 from bovine faeces

AIMS: To compare the sensitivity of two pre-enrichment broth media prior to immunomagnetic separation for the isolation of Escherichia coli O157 from cattle faeces. METHODS AND RESULTS: One-gram portions of 721 cattle faeces collected from 43 farms were pre-enriched in buffered peptone water containing vancomycin, cefixime and cefsulodin (BPW-VCC) and buffered peptone water without additives (BPW-WOA), respectively. A total of 137 samples were positive for E. coli O157: 127 pre-enriched with BPW-WOA and 89 pre-enriched in BPW-VCC. Representative isolates were tested for phage type, verotoxin and eae (E. coli attaching and effacing) gene sequences, resulting in the recognition of eight different types. All the E. coli O157 types recognized were isolated by both methods except for three different strains, each of which were isolated only on a single occasion: two by BPW-WOA and another by BPW-VCC. CONCLUSIONS: The results clearly demonstrate, under the conditions of this study, that BPW without antibiotics was the superior pre-enrichment medium for the isolation of E. coli O157 from cattle faeces. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of BPW-WOA in preference to BPW-VCC for the isolation of E. coli O157 from cattle faeces in future research and outbreak studies should lead to a higher number of positive isolates.     

Recruitment of protein phosphatase 1 to the nuclear envelope by A-kinase anchoring protein AKAP149 is a prerequisite for nuclear lamina assembly

Subcellular targeting of cAMP-dependent protein kinase (protein kinase A [PKA]) and of type 1 protein phosphatase (PP1) is believed to enhance the specificity of these enzymes. We report that in addition to anchoring PKA, A-kinase anchoring protein AKAP149 recruits PP1 at the nuclear envelope (NE) upon somatic nuclear reformation in vitro, and that PP1 targeting to the NE is a prerequisite for assembly of B-type lamins. AKAP149 is an integral membrane protein of the endoplasmic reticulum/NE network. The PP1-binding domain of AKAP149 was identified as K(153)GVLF(157). PP1 binds immobilized AKAP149 in vitro and coprecipitates with AKAP149 from purified NE extracts. Affinity isolation of PP1 from solubilized NEs copurifies AKAP149. Upon reassembly of somatic nuclei in interphase extract, PP1 is targeted to the NE. Targeting is inhibited by a peptide containing the PP1-binding domain of AKAP149, abolished in nuclei assembled with membranes immunodepleted of AKAP149, and restored after reincorporation of AKAP149 into nuclear membranes. B-type lamins do not assemble into a lamina when NE targeting of PP1 is abolished, and is rescued upon recruitment of PP1 to the NE. We propose that kinase and phosphatase anchoring at the NE by AKAP149 plays in a role in modulating nuclear reassembly at the end of mitosis.     

稻恶苗菌异核体三种核型核DNA的RAPD分析

利用RAPD技术,对稻恶苗菌TJ47-26异核体的紫、红、白三种核型(同核体)核DNA进行比较,用80种10bp长的随机引物进行DNA扩增,共检测了基因组中近400个位点,未发现三种核型的基因组间有明显差异,表明紫、红、白三种核型的DNA碱基序列高度相似。由此推测TJ47-26异核体的三种不同核型是来自同一个核的变异体。     

枸杞体细胞胚发生过程中Ag~ 对痕量金属离子吸收的影响

以药用植物宁夏枸杞愈伤组织为材料,离体培养诱导体细胞胚发生。采用多重示踪剂和γ射线能谱分析法研究不同浓度AgNO_3处理的枸杞体细胞胚发生过程中对多种痕量金属元素离子的吸收。结果表明:(1)当AgNO_3的浓度小于50mg/L时,随着AgNO_3浓度的增加,多种痕量金属离子的吸收率也随之增加,而超过此浓度后,对多种痕量金属离子的吸收影响不同。Ag~ 对痕量金属离子的吸收有协同,拮抗或竞争的作用。(2)适当浓度的AgNO_3对细胞分化及体细胞胚发生有促进作用。当AgNO_3的浓度小于50mg/L时,随着AgNO_3浓度的增加,体细胞胚的发生频率随之增加。Ag~ 对枸杞体细胞胚发生表现促进作用,当AgNO_3的浓度为50mg/L时,可大大提高愈伤组织中体细胞胚发生,是对照(不加AgNO_3)组的3倍左右。而超过此浓度后,Ag~ 对枸杞体细胞胚发生表现毒害作用,体细胞胚的发生受到明显抑制。     

Evidence for the existence of a nucleolar skeleton attached to the pore complex-lamina in human fibroblasts

This work deals with the types of nuclear skeletal structures obtained from human fibroblast nuclei isolated by different procedures. It is confirmed that, in somatic vertebrate cells, the pore complex-lamina is always observed, whereas the presence of internal nucleolar and extranucleolar residual structures depends upon the method of nuclear isolation used. Furthermore, the results reported here argue for the existence of a nucleolar skeleton different from the nucleolar matrix often observed in different cell types by other investigators. The conditions of nuclear isolation which allow us to visualize this nucleolar skeleton without any other internal residual structures are described. The attachment of the nucleolar skeleton to the lamina suggested by the present data is considered in relation to the in situ position of nucleoli near the nuclear envelope.