Wheat mutants showing altered adult plant disease resistance

Wheat mutants, selected on an altered resistance phenotype to Puccinia striiformis f.sp. tritici, the causal agent of yellow rust, were assessed in greenhouse tests to determine the growth stage at which the mutant phenotype was expressed and which components of yellow rust resistance were altered. Four of the mutants showed reduced levels of yellow rust infection, while three exhibited increased levels of infection. The infection characters altered were those seen after the establishment of hyphal growth, i.e. days after inoculation to sporulation, the percentage of inoculated leaf tissue producing sporulating colonies and the number of spores produced per cm² of inoculated leaf tissue. The altered resistance phenotype was developmentally regulated in all of the mutants. The changes seen in the infection characters differed for each mutant, suggesting that different genes may have been altered. An altered resistance phenotype to brown rust and/or powdery mildew was also seen for some of the yellow rust mutants. This revised version was published online in August 2006 with corrections to the Cover Date.     

高毒力大丽轮枝菌VDG1特异分泌蛋白HSSP致病相关功能分析

以棉花高毒力大丽轮枝菌(Verticillium dahliae Kleb.)菌株VDG1和低毒力大丽轮枝菌菌株VDG2基因组测序数据为基础,通过比较基因组学和分泌蛋白预测发现VDG1中1个特异分泌蛋白,将其命名为HSSP。通过PEG介导的原生质体转化方法获得了该基因的敲除突变株ΔHSSP,并以木糖、淀粉、纤维素、果胶、木聚糖和半乳糖为底物模拟分析了其降解细胞壁组分的能力,发现该突变体的果胶酶、木聚糖酶和半乳糖酶活力较野生型显著降低;通过测定菌丝产量和孢子产量,发现HSSP基因对菌丝和孢子的形成均有重要作用;通过定量蘸根接种法在感病棉种军棉1号上测定致病力,发现HSSP基因敲除突变体的致病力与野生型相比明显减弱。上述结果说明HSSP作为高毒力大丽轮枝菌菌株VDG1中1个特异的分泌蛋白在对寄主棉花的致病过程中发挥了重要作用。     

肝螺杆菌CdtB基因缺失株的构建与验证

试验旨在敲除肝螺杆菌（Helicobacter hepaticus,H.hepaticus）的CdtB基因,采用pcDNA质粒构建自杀质粒,用于敲除肝螺杆菌ATCC 51449菌株CdtB基因。根据同源重组原理,通过电击转化方法将质粒转入肝螺杆菌构建肝螺杆菌CdtB基因缺失株（ΔCdtB）。采用PCR及测序技术对ΔCdtB缺失株进行鉴定。采用生化试验、生长曲线测定检测ΔCdtB缺失株与野生型肝螺杆菌区别。结果显示,获得以质粒pcDNA构建的同源重组敲除质粒pcDNA-ΔCdtB,电击转化肝螺杆菌后经过传代筛选可获得氯霉素阳性转化子。缺失株替换片段的PCR产物为964 bp,大于野生型肝螺杆菌对应产物884 bp,测序结果表明,氯霉素抗性基因已替换了CdtB基因。比较发现,ΔCdtB缺失株与野生型菌株的尿素酶试验结果均呈阳性,并且在生长速率方面与普通血琼脂平板上无显著差异,但最终缺失株在含氯霉素的血琼脂平板上生长更多。除此之外,将获得的缺失株持续传代并进行PCR鉴定未发现回复突变,有较好的遗传稳定性。综上所述,本研究构建的基因敲除质粒pcDNA-ΔCdtB实现了对肝螺杆菌中目标基因的敲除,为肝螺杆菌基因功能研究和致病因子的发掘提供了有效的基因操作工具。     

Status and Perspectives on the Researches of Rice Glutelin Mutants

Rice (Oryza sativa L. ) is one of the model plants for genomics research. As the raising offunctional rice breeding for special usage, glutelin mutants play a more and more important role in the func-tional rice breeding as well as eukaryotic gene expression and regulation research materials. For example, therice cultivar special for the patients suffering from kidney disease and diabetes could be developed from the riceglutelin mutants. In this paper, current researches on characterization, mutation mechanism and breeding us-age of various rice glutelin mutants, especially the low glutelin content cultivars, were all discussed with per-spectives on the trends of the glutelin mutant researches in the era of post-genomics.     

过表达玉米ZmSC1基因增强转基因拟南芥的耐盐性

旨在从玉米中克隆耐盐相关基因ZmSC1,分析其分子特征并在拟南芥中研究其耐盐性的生物学功能。以玉米B73为试验材料,克隆ZmSC1全长序列,与其他物种进行同源性比对,解析其盐诱导表达模式和亚细胞定位情况,将ZmSC1转入到拟南芥突变体atsc和野生型中,观察在盐处理下种子萌发率和主根根长情况,利用荧光定量PCR分析相关逆境（140 mmol·L^-1胁迫）基因的表达量。结果表明,ZmSC1基因全长为423 bp,编码141个氨基酸,ZmSC1和小麦、拟南芥中已知的TaSC1、AtSC1具有较高保守性。烟草细胞瞬时表达、玉米原生质体亚细胞定位研究表明ZmSC1定位于细胞膜中。生物学功能研究发现在140 mmol·L^-1的NaCl的盐处理下,相比较于拟南芥突变体,回补植株拟南芥的种子萌发率和主根根长得到了明显改善,这说明ZmSC1基因可以回补拟南芥同源基因AtSC1突变体植株在盐胁迫下的表型。ZmSC1基因过表达拟南芥植株的种子萌发率和主根根长也显著高于野生型植株。荧光定量PCR结果显示相较于野生型,在过表达植株中AtRD29A、AtSOS2、AtSOS3、AtCDPK1等胁迫相关基因的表达量也明显增强。研究结果表明TaSC1、AtSC1的同源基因ZmSC1对提高拟南芥的耐盐性具有重要作用。     

棉花黄萎病菌插入突变体库的构建及致病相关基因DVK1的克隆与鉴定

 利用ATMT（Agrobacterium tumefaciens-mediated transformation）技术自建Verticillium dahliae强致病力落叶型菌株T-DNA插入突变体库,共获得5000个转化子;随机挑选1000个转化子进行致病力鉴定,筛选获得5株致病力衰退的突变体。挑选致病力下降最为明显的突变体d1,通过TAIL-PCR技术最终获得一段长3237 bp的DVK1全长cDNA序列,编码1079个氨基酸的蛋白。基于DNA序列比对发现,DVK1基因含有2个内含子,分别为52 bp和36 bp。进一步比对发现T-DNA插入到DVK1基因启始密码上游740 bp的启动子中。通过遗传互补实验d1 恢复了致病性,进一步证明DVK1基因与黄萎菌致病性相关。     

Flower-Colour Polymorphism in Lupinus pilosus in Israel

Four rare flower-colour mutants were detected in natural populations of Lupinus pilosus in Israel. The breeding systems and genetic bases of three oif them were examined. Mutants were tagged in wild populations and seed obtained upon open pollination was collected and grown in a, net house which was insect-proof. Mutant progenies were used in crosses with each other and with wild-type plants. Our results indicate the possibility that all three mutants are simple Mendelian recessives to the wild-type deep-violet colour and that in L. pilosus this colour is not monogenic but determined by at least two loci. All these mutants show a high degree of selling when open-pollinated in nature.     

诱发籼稻早熟同型系的研究 Ⅲ.早熟同型系的发生与不同系谱亲缘的关系

辐射诱发早熟同型系的发生率与水稻品种亲缘有密切关系。同一亲缘或组合衍生的品种早熟同型系发生率基本相似 ;不同亲缘品种早熟同型系的发生率不一 ,有强、弱和一般 3种类型。一般亲缘差异较大的亲本杂交后代早熟同型系发生率比亲本亲缘差异较小杂交后代高。研究表明 :IR8(Peta×DGWG)、IR1 2 7(Cpslo×Sigadis)以及IR2 4(IR8×IR1 2 7)衍生的品种早熟同型系发生率都较高 ,属于诱发早熟同型系发生的优势系谱     

农杆菌介导的芒果胶孢炭疽菌遗传转化及致病性缺陷突变体的筛选

构建含潮霉素抗性基因和GFP基因的双元载体pCAMBgfp,以其为转化载体用农杆菌EHA105介导的方法对芒果胶孢炭疽菌Cg-8菌株进行遗传转化,以潮霉素抗性和GFP荧光来筛选阳性转化子。然后,通过离体接种芒果叶片和果实观察病斑大小筛选致病性缺陷转化子。结果获得500个阳性转化子,转化效率为平均106个孢子可获得400个左右同时具有潮霉素抗性和GFP荧光的阳性转化子,且其经多次在无潮霉素的培养基上传代后能得到稳定遗传。随机挑选其中13个突变体进行PCR检测,均可扩增出潮霉素基因目的条带,致病性分析从中筛选得到8个致病性减弱或缺失突变体。     

苹果树腐烂病菌Zn(Ⅱ)2Cys6类转录因子Vmzctf-07的鉴定及其功能分析

转录因子是真菌生命过程中重要的调控因子,探究转录因子在苹果树腐烂病菌Valsa mali致病过程中的作用,能够为解析病菌致病机理奠定基础。通过构建Vmzctf-07基因的酵母单杂交和亚细胞定位表达载体,进行转录激活域验证和亚细胞定位分析;通过创制Vmzctf-07基因缺失突变体和回补菌株,进行生长速率、非生物胁迫和致病力测定。结果表明,Vmzctf-07具有转录激活功能,转录激活域可能存在于down区段（aa 1 455~aa 1 644）。Vmzctf-07定位于细胞核中,符合转录因子典型特征。Vmzctf-07敲除突变体的生长速率和致病力均显著下降,同时该基因参与病菌响应渗透胁迫和氧化胁迫。综上,Vmzctf-07具有转录激活功能和核定位信号,且参与调节V.mali的生长发育、非生物胁迫和致病过程。