对CIMMYT小麦品种抗白粉病的鉴定简报

1999～2001年连续2年,对 CIMMYT 提供的165份小麦品种进行成株期田间人工诱发抗白粉病鉴定。鉴定病圃设在本院试验农场,面积为200m~2。人工诱发鉴定圃的接种菌源,以上年度采集并保存的当地白粉病菌菌株,先在温室的感病品种8017-2上繁殖,2月下旬将带有大量分生孢子堆的病株移植到病圃的二个诱发行中间,每距33cm 种植2株病株。由于病株上有大量分生孢子,使诱发行麦株迅速受侵染且严重发病,形成充分接种菌源,保证对供鉴品种的重复侵染,以诱发供试小麦材料成株期感染发病。     

白粉菌侵染对小麦叶片显微及超微结构的影响

通过半薄及超薄切片,比较了正常和受白粉菌感染的小麦叶片细胞的显微及超微结构的差异。观察结果发现：（1）受感染小麦叶肉细胞的细胞壁上局部沉积大量团状电子致密颗粒;（2）叶绿体形状由原来的椭圆形转变成圆形,叶绿体膜破裂;类囊体膨大,基粒片层排列疏松,同时,叶绿体内嗜饿性颗粒数量增加;（3）线粒体膜解体,内含物分散到了细胞质中。     

Differential regulation of wheat quinone reductases in response to powdery mildew infection

At least two types of quinone reductases are present in plants: (1) the ζ-crystallin-like quinone reductases (QR1, EC 1.6.5.5) that catalyze the univalent reduction of quinones to semiquinone radicals, and (2) the DT-diaphorase-like quinone reductases (QR2, EC 1.6.99.2) that catalyze the divalent reduction of quinones to hydroquinones. QR2s protect cells from oxidative stress by making the quinones available for conjugation, thereby releasing them from the superoxide-generating one electron redox cycling, catalyzed by QR1s. Two genes, putatively encoding a QR1 and a QR2, respectively, were isolated from an expressed sequence tag collection derived from the epidermis of a diploid wheat Triticum monococcum L. 24 h after inoculation with the powdery mildew fungus Blumeria graminis (DC) EO Speer f. sp. tritici Em. Marchal. Northern analysis and tissue-specific RT-PCR showed that TmQR1 was repressed while TmQR2 was induced in the epidermis during powdery mildew infection. Heterologous expression of TmQR2 in Escherichia coli confirmed that the gene encoded a functional, dicumarol-inhibitable QR2 that could use either NADH or NADPH as an electron donor. The localization of dicumarol-inhibitable QR2 activity around powdery mildew infection sites was accomplished using a histochemical technique, based on tetrazolium dye reduction.     

Genome-wide analysis of long non-coding RNAs in Pichia pastoris during stress by RNA sequencing

Long non-coding RNAs play significant roles in many biological processes. The roles of lncRNAs in Pichia pastoris remain unclear. In this work, we focused on the identification of lncRNAs in P. pastoris and exploration of their potential roles in stress response to PLA₂ overexpression and methanol induction. By strand specific RNA sequencing, 208 novel long non-coding RNAs were identified and analyzed. Bioinformatic analysis showed potential trans-target genes and cis-regulated genes of 39 differential lncRNAs. Functional annotation and sequence motif analysis indicated that lncRNAs participate in pathways related to methanol degradation and production of the recombinant protein. The differential expression of lncRNAs was validated by qRT-PCR. Lastly, the potential functions of three lncRNAs were evaluated by knockdown of their expression and analysis of the expression levels of target genes. Our study identifies novel lncRNAs in P. pastoris induced during use as a bioreactor, facilitating future functional research.     

Diverse set of microRNAs are responsive to powdery mildew infection and heat stress in wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

Background  

MicroRNAs (miRNAs) are a class of small non-coding regulatory RNAs that regulate gene expression by guiding target mRNA cleavage or translational inhibition. MiRNAs can have large-scale regulatory effects on development and stress response in plants.     

小麦抗白粉病基因Pm23的RAPD标记

小麦抗白粉病基因Pm23对世界上很多麦区流行的白粉病表现高抗或免疫.本研究以Pm23和Chancellor为抗感亲本,用集群分离分析法对抗性基因Pm23进行了RAPD分析,从320个十碱基随机引物中筛选到一个与Pm23紧密连锁的相引相标记OPE051100. 对F2分离群体进行RAPD分析表明,该标记与Pm23基因之间的连锁距离为10.65±3.25 cM.该标记可以有效用于小麦育种分子标记辅助选择中.     

抗白粉病小麦——中间偃麦草异附加系的细胞学和RAPD鉴定

利用细胞学和ＲＡＰＤ技术,对从小麦与中间偃麦草杂种后代中选育的抗白粉病异附加系ＤＡＬ６６进行了鉴定。结果证明ＤＡＬ６６根尖细胞染色体数为４４,花粉母细胞减数第一分裂中期（ＰＭＣ ＭＩ）杂色体模型为２ｎ＝２２Ⅱ。对ＤＡＬ６６及其双亲进行ＲＡＰＤ分析,从４０个随机引物中筛选出１个特异引物（ＯＰＥ－０２）能够稳定地扩增出特异带型。     

甜瓜S-腺苷甲硫氨酸脱羧酶基因的克隆及白粉病诱导表达分析

为探讨S-腺苷甲硫氨酸脱羧酶(S-adenosylmethionine decarboxylase,SAMDC)基因在甜瓜抵御白粉病菌中的作用,根据已知EST序列和甜瓜基因组数据库,在甜瓜抗白粉病品种‘Yuntian930’中克隆获得该基因的全长编码序列,命名为CmSAMDC(GenBank登录号为KF151861)。生物信息学分析表明,CmSAMDC的主开放阅读框(mORF)长1 095 bp,编码364个氨基酸,预测分子量为40 kDa。聚类分析表明,CmSAMDC预测蛋白与黄瓜和四季橘中该蛋白的同源关系最近,并与其他双子叶植物聚为一类。原核表达分析表明,CmSAMDC以融合蛋白形式表达,相对分子量约为40 kDa,与预测一致。实时定量表达分析表明,CmSAMDC基因受白粉病诱导表达,在接种后48 h表达量达到峰值,为接种前的7倍,并且在甜瓜的根、茎、叶、卷须中均有表达。结果提示该基因可能参与了甜瓜的抗白粉病反应。     

Genetic mapping of the powdery mildew resistance gene Pm6 in wheat by RFLP analysis

Pm6 in bread wheat (Triticum aestivum L.), which was transferred from Triticum. timopheevii L., is a gene conferring resistance to the powdery mildew disease caused by Erysiphe graminis f. sp. tritici. Six near-isogenic lines ( NILs ) of Pm6 in a cultivar ’Prins’ background were analyzed to map this gene using restriction fragment length polymorphism (RFLP). Each of the six NILs possessed a T. timopheevii-derived segment, varying in length, and associated with powdery mildew resistance. Lines IGV1–465 (FAO163b/ 7*Prins) and IGV1–467 (Idaed 59B/7*Prins) had the shortest introgressed segments, which were detected only by DNA probes BCD135 and PSR934, respectively. The polymorphic loci detected by both probes were mapped to the long arm of chromosome 2B. Lines IGV1–458 (CI13250/7*Prins) and IGV1–456 (CI12559/8*Prins) contained the longest T. timopheevii segments involving both arms of donor chromosome 2G across the centromere. All these introgressed segments had an overlapping region flanked by the loci xpsr934 and xbcd135 on 2BL. Thus, Pm6 was located in this region since the powdery mildew resistance in all the NILs resulted from the introgressed fragments. Using the F₂ mapping population from a cross of IGV1–463 (PI170914/7*Prins)×Prins, Pm6 was shown to be closely linked to the loci xbcd135 and xbcd266 at a genetic distance of 1.6 cM and 4.8 cM, respectively. BCD135 was successfully used in detecting the presence of Pm6 in different genetic backgrounds. Received: 29 June 1999 / Accepted: 6 July 1999     

Transgenic wheat progeny resistant to powdery mildew generated by Agrobacterium inoculum to the basal portion of wheat seedling

To improve the transformation efficiency of wheat (Triticum aestivum L.) mediated by Agrobacterium tumefaciens, we explored the possibility of employing the basal portion of wheat seedling (shoot apical meristem) as the explants. Three genotypes of wheat were transformed by A. tumefaciens carrying β-1, 3-glucanase gene. After vernalization, the seeds to be transformed were germinated. When these seedlings grew up to 2∼5 cm, their coleoptile and half of the cotyledon were cut out, and the basal portions were infected by A. tumefaciens. A total 27 T₀ transgenic plants were obtained, and the average transformation efficiency was as high as 9.82%. Evident segregation occurred in some of the T₁ plants, as was indicated by PCR and Southern blotting analysis. Investigation of the T₂ plants revealed that some transformed plants had higher resistance to powdery mildew than the controls. Northern blotting revealed that β-1, 3-glucanase gene was normally expressed in the T₂ plants, which showed an increased resistance to powdery mildew. The results above indicate that the exogenous gene has been successfully integrated into the genome of wheat, transmitted and expressed in the transgenic progeny. From all the results above, it can be concluded that Agrobacterium inoculum to the basal portion of wheat seedling is a highly efficient and dependable transformation method. It can be developed into a practicable method for transfer of target gene into wheat.Tong-Jin Zhao and Shuang-Yi Zhao contributed equally to this paper.     

The expression of resistance to powdery mildew infection in winter wheat cultivars. I. Seedling resistance

Detached seedling leaf tests were used to assign 18 winter wheat cultivars to three wheat mildew resistance (WMR) groups on the basis of identified race-specific resistance characters. The same cultivars were exposed in mobile seedling nurseries from 1976 to 1979. Infection by mildew populations containing matching pathogenicity characters was used to recognise background seedling resistance in three cultivars: Maris Freeman, Maris Huntsman and Sentry. The value of background seedling resistance is discussed.     

抗白粉病小偃麦异代换系的细胞学和RAPD鉴定

利用细胞学和RAPD方法,对从长穗偃麦草与普通小麦复合杂交后代中选育的抗白粉病小麦种质系山农87074－526和山农87074－551进行了鉴定。结果表明,两种质系的根尖细胞染色体数目均为2n=42,花粉母细胞减数分裂中期I（PMC MI）染色体构型为2n=21Ⅱ;二者杂交F1 PMC MI染色体构型亦为2n=21Ⅱ,两种质系分别与小麦中国春的杂种F1 PMC MI染色体构型均为2n=20Ⅱ 2I,说明两种质系为相同的双体异代系。在苗期和成株期两种质系对白粉病15号菌种均表现免疫,其白粉病抗性为显性,并且来自长穗偃麦草,抗白粉病基因位于它们所含的偃麦草染色体上。从80个随机引物中,筛选出2个引物OPE13和OPH15能在两种质系中稳定地扩增出长穗偃麦草亲本的特异DNA片段。     

Identification and molecular mapping of a resistance gene to powdery mildew from the synthetic wheat line M53

Powdery mildew disease caused by Blumeria graminis f. sp. tritici (Bgt) is an economically important disease in wheat worldwide. The identification of germplasms resistant to the disease can not only facilitate the breeding of resistant cultivars, but can also broaden the diversity of resistance genes. The Mexican M53 is a synthetic hexaploid wheat line developed at the International Maize and Wheat Improvement Center (CIMMYT) from the cross between Triticum durum and Aegilops tauschii249. Infection of M53 with 15 different pathogen races revealed that the resistance in M53 was race-dependent and effective against the majority of the tested Bgt races, including the race 15 predominant in the Beijing wheat growing area. Inoculation of the parents of M53 with the race 15 demonstrated that M53 and Ae. tauschii249 were resistant, whereas T. durum was susceptible. The inoculation of three segregating F₂ populations developed from the crosses between M53 and three susceptible Chinese wheat cultivars with the race 15 showed that the resistant gene in M53 segregated in a single dominant manner. Amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) markers were used to map the gene in a segregating F₂ population consisting of 213 lines developed from the cross Wan7107 × M53. Two closely linked AFLP markers, Apm109 and Apm161, were identified to flank the gene with genetic distances of 1.0 cM and 3.0 cM, respectively. The recognized gene was assigned to the long arm of chromosome 5D as determined by three linked SSR markers, Xwmc289b, Xgwm583, and Xgwm292, and by the physical mapping of Apm109 using Chinese Spring nullisomic–tetrasomic and ditelosomic stocks. The resistance gene identified in M53, temporarily designated as Pm-M53, could be used in local wheat-breeding programs to improve powdery mildew resistance.     

Identification and mapping of a new powdery mildew resistance allele in the Chinese wheat landrace Hongyoumai

Powdery mildew (Pm) caused by Blumeria graminis f. sp. tritici (Bgt) is one of the world’s major wheat diseases and results in large grain yield losses. Discovery and utilization of Pm resistance genes constitute the most common strategy for wheat Pm control. Hongyoumai, a wheat landrace from Henan Province in China, has excellent resistance to infection by Bgt. In order to identify the basis of such Pm resistance, a segregating population was submitted to genetic analysis, which showed that Pm resistance in Hongyoumai was conferred by a single recessive resistance gene. This gene was temporarily named pmHYM. Molecular marker analysis, chromosomal location, resistance spectrum analysis, and an allelism test showed that pmHYM was located on the long arm of chromosome 7B (7BL), most likely representing a new recessive resistance gene allelic with Pm5e and mlXBD. By using 90-kb single-nucleotide polymorphism sequences (SNP) in the BLASTn analysis against the wheat 7BL genome sequence, 12 new simple sequence repeat (SSR) markers linked with pmHYM were developed to map pmHYM co-segregating with the marker Xmp1207 and between markers Xmp925 and Xmp1158, at genetic distances of 2.8 and 2.7 cM, respectively. In addition, physical mapping of the markers linked with pmHYM using Chinese Spring deletion lines indicated a location in the 0.86–1.00 bin of 7BL.