宿根矮化病抗感甘蔗品种茎部内生真菌和细菌群落特征分析

通过分析宿根矮化病抗感甘蔗品种蔗茎内生真菌和细菌的群落特征,旨在了解内生真菌和细菌类群在寄主抗病中的作用,从而为构建生态防控宿根矮化病提供理论依据。利用Illumina MiSeq高通量测序技术,比较了宿根矮化病感病品种YT60和抗病品种HoCP95988蔗茎内生真菌及细菌的多样性指数、类群组成和物种差异。感病品种YT60蔗茎内生真菌和细菌的Alpha多样性指数高于抗病品种HoCP95988,具有更多的物种组成。蔗茎内生细菌主要分布在变形菌门(Proteobacteria),子囊菌门(Ascomycota)和担子菌门(Basidiomycota)为内生真菌的优势菌门。抗感病甘蔗品种蔗茎内生真菌和细菌种群组成差异明显,其中内生细菌在浮霉菌门(Planctomycetes)、绿弯菌门(Chloroflexi)、芽单胞菌门(Gemmatimonadetes)和软壁菌门(Tenericutes)中包含的34个属的物种丰度存在显著差异,并且差异物种在感病品种YT60中呈现较高相对丰度。宿根矮化病抗感甘蔗品种茎部内生真菌和细菌多样性均较为丰富,种群组成存在较大差异,差异物种的分类信息为宿根矮化病生...     

甘蔗宿根矮化病研究进展

综述了近年来甘蔗宿根矮化病的检测、传播、防治及其致病菌Leifsonia xyli subsp.xyli的分类归属和致病机理等方面的研究进展,并探讨了研究策略.     

甘蔗主要病害抗病遗传育种研究进展

甘蔗生产长期受到病害的严重影响,这些病害不仅降低蔗茎产量和糖分,还影响宿根性,甚至造成品种种性退化,而抗病品种的选育是保证甘蔗生产的有效途径。概述了甘蔗花叶病、黑穗病、宿根矮化病和梢腐病4种主要甘蔗病害在病原物检测、种质筛选和基因工程技术等抗病育种工作方面的研究进展。     

甘蔗栽培原种宿根蔗综合性状分析

对50份甘蔗栽培原种的宿根萌发率、分蘖率、株高、茎径、锤度、蔗糖分及自然条件下感病情况进行综合评价分析。结果表明：50份材料宿根萌发率为7%~184%,分蘖率为0~360%,宿根蔗中有效茎数、株高、茎径、锤度和蔗糖分值最高的材料分别是光泽竹蔗、四川芦蔗、拔地拉、德阳大叶子和28NG251。以各性状的前10份优异材料进行评价,筛选出21份含有2个以上优良性状的材料,含3个优良性状的有11份材料,含4个优良性状的有1份材料,其中以热带种14NG124的综合性状最佳。从感病率来看,中国种感染花叶病和黑穗病的比率高于热带种和印度种,其中德阳大叶子、广西竹蔗和文山蔗同时感染花叶病和黑穗病。     

甘蔗宿根萌发期內源激素变化与其宿根性的关系

采用酶联免疫法和茎基部取样的方式,以强宿根甘蔗品种‘新台糖22号’为对照,通过田间试验研究了同一组合(‘粤糖97-20’×ROC25)杂交后代分离产生的强宿根甘蔗品(种)系(‘云蔗06-407’、‘云蔗06-415’)和弱宿根甘蔗品(种)系(‘云蔗06-408’、‘云蔗06-416’)宿根萌发期内源激素含量的变化与甘蔗宿根性的关系。结果显示:(1)宿根萌发过程中,强宿根与弱宿根甘蔗茎基部IAA含量、ZR含量和ZR/IAA比值的变化波动大,但差异不显著。(2)强宿根与弱宿根甘蔗茎基部GA3和ABA含量、IAA/ABA和GA3/ABA比值的变化差异较大,其中ABA含量、IAA/ABA和GA3/ABA比值的差异显著,是与甘蔗宿根性关系密切的3个重要参数。(3)强宿根甘蔗ABA含量高于弱宿根甘蔗,而其IAA/ABA和GA3/ABA比值小于弱宿根甘蔗,且IAA/ABA和GA3/ABA比值随甘蔗宿根性的减弱而增大。研究认为,在宿根萌发期,甘蔗宿根性越强,ABA含量越高,IAA/ABA和GA3/ABA比值越小;而这3个重要参数均与ABA有关,进行甘蔗宿根性评价应着重参考ABA含量,同时结合IAA/ABA和GA3/ABA比值再进行深入分析鉴定。     

中国基因组学研究进展与发展态势

20 世纪 90 年代初,以完成人类基因组全序列测定和注释为核心任务的人类基因组计划在美国的领导下兴起．自1999年中国加入人类基因组计划到现在的10年时间里,中国基因组学得到了快速的发展,建立了先进的基因组学技术平台,并出色完成了多项重大基因组科学研究项目,对我国生命科学各个领域的发展产生了重要影响．结合我国基因组学研究现状,《中国科学C辑·生命科学》(Sci China Ser C-Life Sci) 2009年第1期发表了中国基因组学专题,综述了基因组测序、分型,功能基因检测技术和生物信息学分析技术,以及肝癌、免疫和环境与工业微生物的基因组学研究等方面的研究工作．     

流感嗜血杆菌的基因组学研究进展

流感嗜血杆菌Rd型是第一个被测序的细菌.其基因组大小为1830137bp,A含量为31%,C为19%,G为19%,T为31%,GC含量为38%,TIGR确认了1473个具有重要功能的基因.基因组序列中包含有复制子、核糖体启动子、毒力基因、DNA转运系统、调节子等结构.流感嗜血杆菌的基因组研究必将对寻找疾病相关基因和抗原基因,筛选药物作用靶位,研制特异的诊断试剂、药物和疫苗具有十分重要的科学意义.对流感嗜血杆菌的基因组学研究进展进行了综述.     

灵长类比较基因组学的研究进展

随着人类和黑猩猩全基因组测序工作宣布完成,以及其他灵长类基因组测序工作的逐步开展,目前已经积累了大量的灵长类基因组数据,一个崭新的研究领域——灵长类比较基因组学应运而生。该文主要通过对人类和其他非人灵长类系统关系和基因组结构的比较,从系统进化、基因组结构和基因表达调控等方面评述该领域的研究进展,阐述人类、黑猩猩与其他非人灵长类之间的主要生物学差异,揭示人类进化的生物学机制。     

高质量甘蔗基因组DNA的简便快速提取方法研究

甘蔗是世界上重要的糖料作物和能源作物。目前,甘蔗分子生物学研究已成为甘蔗研究的热点之一。基因组DNA的提取是进行甘蔗分子生物学研究的基础。本研究设计含一系列SDS浓度的提取液,同时设加液氮和不加液氮研磨的对比试验,提取甘蔗不同部位叶片的基因组DNA并进行产量和纯度检测以及分子生物学分析。结果表明,所有提取液提取的甘蔗基因组DNA纯度均很高,A260/A280在1.8-2.0之间,A260/A230大于2,但提取液I(0.75%SDS)提取的甘蔗基因组DNA产量较低;加液氮与否对甘蔗基因组DNA的提取产量和纯度没有影响;以提取的甘蔗基因组DNA为模板,分别用一对扩增SPS(蔗糖磷酸合成酶)基因部分片段的引物和一对ISSR引物进行PCR扩增,所有DNA均能扩增出预期的条带;用不同的限制性内切酶对所提取的甘蔗基因组DNA进行酶切,所有DNA样品均能完全酶切。本研究得出最佳甘蔗基因组DNA提取方法如下:磨碎甘蔗叶片后,加DNA提取液(SDS:1.5%;Tris:100 mM;EDTA:20 mM;NaCl:500 mM)于65℃裂解30 min,经酚∶氯仿和氯仿各抽提一次,可获得高产量高质量的甘蔗基因组DNA,能满足后续分子生物学研究的要求。     

基于高精度转录组测序技术挖掘甘蔗抗梢腐病相关的NBS-LRR家族基因

甘蔗(Saccharum officinarum)是中国乃至世界第一大糖料作物,也是非常重要的生物能源作物。甘蔗在大田生长过程中经常遭遇病原菌侵入而诱发甘蔗病害,严重时会造成产量下降、品质受损,在广西蔗区常年大面积种植甘蔗,梢腐病的发生极为普遍,也常因此造成严重的经济损失。本研究以具有易感梢腐病特征的现代栽培种‘中蔗1号’为实验材料,采用一种更高精度的转录组测序新技术,在全基因组水平鉴定到含有NB-ARC结构域的NBS抗病基因(含等位基因)共2 324个,其中在‘中蔗1号’梢腐病病株与健康蔗株中存在差异表达的基因有101个。进一步对这些基因的结构域进行注释分析,确定了36个NBS-LRR类抗病基因。推测这些基因可能参与甘蔗对梢腐病致病菌的抵御以及病害侵染甘蔗后的一系列防御反应。     

Incidence and Severity of Leifsonia xyli subsp. xyli Infection of Sugarcane in Sao Paulo State,Brazil

Sugarcane covers 8.53 million hectares with production of 596.63 million tonnes in Brazil. Despite its importance, little information is available on ratoon stunt (RSD), caused by Leifsonia xyli subsp xyli (Lxx). Our objective was to examine the incidence and severity of Lxx among sugarcane cultivars in 2009, 2010 and 2011. Sap from 100 stalks from each field was sent for a routine RSD analyses that allowed examination of Lxx incidence. The presence of bacterium was checked by dot blot enzyme immunoassay to detect its presence and relative concentration. Analyses of 187 fields from 35 cultivars in 2009, 166 fields from 33 cultivars in 2010 and 221 fields from 30 cultivars in 2011 found Lxx incidence of 23.6% of fields of 23 cultivars in 2009, 27.1% of fields of 15 cultivars in 2010 and 25.8% of fields of 15 cultivars in 2011. RB867515, the major cultivar in Sao Paulo, had within‐field incidence of up to 70% in 2009, 48% in 2010 and 88% in 2011. Highest incidence and populations of Lxx infection were found for cvs RB867515, RB855453, SP81‐3250, RB855536 and RB92579, demonstrating their susceptibility to RSD.     

Establishment of a functional genomics platform for Leifsonia xyli subsp. xyli

Leifsonia xyli subsp. xyli, the causal agent of ratoon stunting disease in sugarcane, is a xylem-limited, nutritionally fastidious, slow growing, gram-positive coryneform bacterium. Because of the difficulties in growing this bacterium in pure culture, little is known about the molecular mechanisms of pathogenesis. Currently, the genome sequence of L. xyli subsp. xyli is being completed by the Agronomical and Environmental Genomes group from the Organization for Nucleotide Sequencing and Analysis in Brazil. To complement this work, we produced 712 Lxx::Tn4431 transposon mutants and sequenced flanking regions from 383 of these, using a rapid polymerase chain reaction-based approach. Tn4431 insertions appeared to be widespread throughout the L. xyli subsp. xyli genome; however, there were regions that had significantly higher concentrations of insertions. The Tn4431 mutant library was screened for individuals unable to colonize sugarcane, and one noncolonizing mutant was found. The mutant contained a transposon insertion disrupting two open reading frames (ORF), one of which had homology to an integral membrane protein from Mycobacterium leprae. Sequencing of the surrounding regions revealed two operons, pro and cyd, both of which are believed to play roles in disease. Complementation studies were carried out using the noncolonizing Lxx::Tn4431 mutant. The noncolonizing mutant was transformed with a cosmid containing 40 kbp of wild-type sequence, which included the two ORF disrupted in the mutant, and several transformants were subsequently able to colonize sugarcane. However, analysis of each of these transformants, before and after colonization, suggests that they have all undergone various recombinant events, obscuring the roles of these ORF in L. xyli subsp. xyli pathogenesis.     

The genome sequence of the gram-positive sugarcane pathogen Leifsonia xyli subsp. xyli

The genome sequence of Leifsonia xyli subsp. xyli, which causes ratoon stunting disease and affects sugarcane worldwide, was determined. The single circular chromosome of Leifsonia xyli subsp. xyli CTCB07 was 2.6 Mb in length with a GC content of 68% and 2,044 predicted open reading frames. The analysis also revealed 307 predicted pseudogenes, which is more than any bacterial plant pathogen sequenced to date. Many of these pseudogenes, if functional, would likely be involved in the degradation of plant heteropolysaccharides, uptake of free sugars, and synthesis of amino acids. Although L. xyli subsp. xyli has only been identified colonizing the xylem vessels of sugarcane, the numbers of predicted regulatory genes and sugar transporters are similar to those in free-living organisms. Some of the predicted pathogenicity genes appear to have been acquired by lateral transfer and include genes for cellulase, pectinase, wilt-inducing protein, lysozyme, and desaturase. The presence of the latter may contribute to stunting, since it is likely involved in the synthesis of abscisic acid, a hormone that arrests growth. Our findings are consistent with the nutritionally fastidious behavior exhibited by L. xyli subsp. xyli and suggest an ongoing adaptation to the restricted ecological niche it inhabits.     

Transformation and transposon mutagenesis of Leifsonia xyli subsp. xyli, causal organism of ratoon stunting disease of sugarcane

Conditions have been developed for genetic transformation and insertional mutagenesis in Leifsonia xyli subsp. xyli (Lxx), the causal organism of ratoon stunting disease (RSD), one of the most damaging and intractable diseases of sugarcane internationally. Transformation frequencies ranged from 1 to 10 colony forming units (CFU)/microg of plasmid DNA using Clavibacter/Escherichia coli shuttle vectors pCG188, pDM302, and pDM306 and ranged from 50 to 500 CFU/microg using cosmid cloning vectors pLAFR3 and pLAFR5-km. The transformation/transposition frequency was 0 to 70 CFU/microg of DNA, using suicide vectors pUCD623 and pSUP2021 containing transposable elements Tn4431 and Tn5, respectively. It was necessary to grow Lxx in media containing 0.1% glycine for electroporation and to amplify large plasmids in a dam-/dcm- E. coli strain and purify the DNA by anion exchange. To keep selection pressure at an optimum, the transformants were grown on nitrocellulose filters (0.2-microm pore size) on media containing the appropriate antibiotics. Transposon Tn4431 containing a promoterless lux operon from Vibrio fischeri and a tetracycline-resistance gene was introduced on the suicide vector pUCD623. All but 1% of the putative transposon mutants produce light, indicating transposition into functional Lxx genes. Southern blot analysis of these transformants indicates predominantly single transposon insertions at unique sites. The cosmid cloning vector pLAFR5-km was stably maintained in Lxx. The development of a transformation and transposon mutagenesis system opens the way for molecular analysis of pathogenicity determinants in Lxx.     

A novel toxin-antitoxin operon talA/B from the Gram-positive bacterium Leifsonia xyli subsp. cynodontis

Here we report a toxin-antitoxin (TA) operon talAB identified from the Gram-positive bacterium Leifsonia xyli subsp. cynodontis. It is shown that talB encodes a broad-host cytotoxin functioning in different Gram-positive bacteria, while talA encodes its antidote. TalA and TalB form different hetero-oligomers in vitro; these hetero-oligomers, but not the antitoxin TalA, strongly bind to the talAB promoter region containing two inverted repeats. This represents a new mechanism of binding the promoter of a TA operon by the toxin and antitoxin complexes.