The VER2 promoter contains repeated sequences and requires vernalization for its activity in winter wheat （Triticum aestivum L.）

Previously, we isolated a vernalization-related gene, VER2, from winter wheat (Triticum aestivum L.) and its expression was restricted in the immature leaves of vernalized wheat seedlings. To further investigate the regulation of VER2 expression and the function of its promoter, we isolated a 41.7 kb genomic clone containing VER2 gene from atransformation-competent artificial chromosome (TAC) library of wheat (Triticum aestivum-Haynaldia villosa). The sequence analysis showed that there were eleven predicted genes in the TAC. The exons of gene 3 corresponded to the cDNA sequence of VER2 gene. Analysis of VER2 promoter structure showed that there were three small repeat sequences divided by two large repeat sequences. The putative response elements, such as abscisic acid response elements (ABRE), MeJA-response elements (Me-JARE), low-temperature response elements (LTR), endosperm expression elements, MYB binding sites and similar elements to GA response elements (GARE), were involved in the VER2 promoter region. Construct containing the VER2 promoter (-5895 to 73) driving GFP reporter gene was bombarded into vernalized or non-verualized immature leaves in wheat. The vernalized immature leaves showed bright green fluorescence after incubation for 24 h, however, the green fluorescence was not observed in the non-vernalization leaves under the same condition. These results suggested that vernalization was essential for the function of VER2 promoter in the immature leaves of winter wheat.     

Development of Triticum aestivum-Leymus racemosus ditelosomic substitution line 7Lr#1S（7A） with resistance to wheat scab and its meiotic behavior analysis

Leymus racemosus is highly resistant to wheat scab （Fusarum head bright）. The transfer of scab resistant gene from L. racemosus to Triticum aestivum is of great significance for broadening the base of wheat resistance. In the present study, the pollen of T. aestivum-L, racemosus monosomic addition line with scab resistance was treated by irradiation with 1200 R ^60Co-γ-rays prior to pollinating to emasculated wheat cv. Mianyang 85-45. Nine plants with a telocentric chromosome 7Lr#1S were observed in M1, and one ditelosomic substitution line 7Lr#1S was selected from selfcrossing progenies and confirmed by chromosome C-banding and GISH. Furthermore, a co-dominant EST-SSR marker CINAU 31 was employed to identify this substitution line. A pair of chromosome 7A of common wheat were found to be replaced by a pair of telocentric chromosome 7Lr#1S, and further investigation showed that chromosome configuration of the substitution line at MI of PMCs after GISH was 17.50Ⅱ^w ＋ 2.19Ⅱ^w ＋ 0.42Ⅱ^7Lr#1S ＋ 1.08 Ⅰ^7Lr#1S ＋ 0.69 Ⅰ^w. Two telocentric chromosomes paired as a bivalent in 59.7% of PMCs. Abnormal chromosome behaviors of telocentric chromosomes were observed in part of PMCs at anaphase I and telophase I, including the moving of two telocentric chromosomes to the same pole, lagging and earlier separation of their sister chromatid. All these abnormal behaviors can be grouped into three distinct types of tetrads according to different numbers of 7Lr#1S in their daughter cells and various micronucleus in some tetrads. However, due to the high transmission frequency of the female and male gametes with a 7Lr#1S, 84% of the selfcrossing progeny plants had ditelosomic substitution. The substitution line showed high resistance to wheat scab in a successive two-year test both in the greenhouse and field; hence, the line will be particularly valuable for alien gene mapping, small fragment translocation induction and telosomic cytological behavior analysis.     

Development of Triticum aestivum-Leymus racemosus ditelosomic substitution line 7Lr#1S(7A) with resistance to wheat scab and its meiotic behavior analysis

Leymus racemosus is highly resistant to wheat scab (Fusarum head bright). The transfer of scab resistant gene from L. racemosus to Triticum aestivum is of great significance for broadening the base of wheat resistance. In the present study, the pollen of T. aestivum-L. racemosus monosomic addition line with scab resistance was treated by irradiation with 1200 R 60Co-γ-rays prior to pollinating to emasculated wheat cv. Mianyang 85-45. Nine plants with a telocentric chromosome 7Lr#1S were observed in M1, and one ditelosomic substitution line 7Lr#1S was selected from selfcrossing progenies and confirmed by chromosome C-banding and GISH. Furthermore, a co-dominant EST-SSR marker CINAU 31 was employed to identify this substitution line. A pair of chromosome 7A of common wheat were found to be replaced by a pair of telocentric chromosome 7Lr#1S, and further investigation showed that chromosome configuration of the substitution line at MI of PMCs after GISH was 17.50○II W 2.19 IIW 0.42II7Lr#1S 1.08 I7Lr#1S 0.69 IW. Two telocentric chromosomes paired as a bivalent in 59.7% of PMCs. Abnormal chromosome behaviors of telocentric chromosomes were observed in part of PMCs at anaphase I and telophase I, including the moving of two telocentric chromosomes to the same pole, lagging and earlier separation of their sister chromatid. All these abnormal behaviors can be grouped into three distinct types of tetrads according to different numbers of 7Lr#1S in their daughter cells and various micronucleus in some tetrads. However, due to the high transmission frequency of the female and male gametes with a 7Lr#1S, 84% of the selfcrossing progeny plants had ditelosomic substitution. The substitution line showed high resistance to wheat scab in a successive two-year test both in the greenhouse and field; hence, the line will be particularly valuable for alien gene mapping, small fragment translocation induction and telosomic cytological behavior analysis.     

Cloning of a down-regulated gene encoding small GTP binding protein in hybrid wheat

Previous studies showed that differential gene expression between wheathybrids and their parents was responsible for the heterosis. To provide an insight into the molecular basis of wheat heterosis, one cDNA, designated TaRab, was identified from the cDNA library of wheat seedling leaves. The sequence comparison in GenBank revealed that TaRab is homologous to a group of genes encoding Rab-GTP binding protein. Semi-quantitative RT-PCR analysis indicated that TaRab was expressed in all plant tissues examined, but at slightly higher level in leaves. Further analysis exhibited that TaRab displayed lower expression in hybrid than in its patents in both roots and leaves, which was in agreement with the original results of suppression subtractive hybridization. TaRab was located on chromosome 7B and C-7DS5-0.36 by in silico mapping. The relationship between differential expression of TaRab and the molecular basis of wheat heterosis was also discussed.     

Cloning and characterization of a glucose 6-phosphate／phosphate translocator from Oryza sativa

Plastids of nongreen tissues import carbon as a source of biosynthetic pathways and energy, and glucose 6-phosphate is the preferred hexose phosphate taken up by nongreen plastids. A cDNA clone encoding glucose 6-phosphate/phosphate translocator (GPT) was isolated from a cDNA library of immature seeds of rice and named as OsGPT. The cDNA has one uninterrupted open reading frame encoding a 42 kDa polypeptide possessing transit poptide consisting of 70 amino acid residues. The OsGPT gene maps on chromosome 8 of rice and is linked to the quantitative trait locus for 1000-grain weight. The expression of OsGPT is mainly re-stricted to heteretrephic tissues. These results suggest that glucose 6-phosphate imported via GPT can be used for starch biosynthesis in rice nongreen plastids     

Polycistronic strategy for cyanobacterial expression vector construction: Co-transcription of a human gene and a selective marker gene

A polycistronic expression vector, pKGA-NTF1, was constructed for the cyanobacterium. Within this vector, the spectinomycin/streptomycin resistance gene (aadA) facilitated the selection of transformants when co-transcribed with favorite genes. A natural glnA gene was selected as the platform to introduce the plasmid into a neutral site of the Synechococcus sp. PCC 7002 chromosome. Function of the vector was demonstrated by the insertion of a modified human Trefoil factor 3 gene (NTF1) to upstream of the aadA gene and by the analyses of the transformed strains. Antibiotics resistance assays showed that the dicistronic expression cassette conferred high spectinomycin resistance to both the E. coli cells and the Synechococcus cells. PCR analysis and Western-blot analysis were carried out to confirm the integration and expression of the NTF1 gene, respectively. Through simple molecular manipulations, the artificial polycistronic structure described here can be conveniently used to express other favorable genes or operons in cyanobacteria, and to study the cyanobacterial gene expression as well.     

Evaluation and characterization of an endosperm-specific sbeⅡa promoter in wheat

Hexaploid wheat (bread wheat) is widely grown all over the world. Wheat grain, which contains high levels of starch and protein located in the endosperm, is being developed as a 揵ioreactor?using the new gene transfor-mation technology[1]. Some new traits (such as virus and insect resistance) have been successfully introduced into key cereal crops using transgene technology, however, there are only a few reports on the studies of transgene expression in wheat because of technical difficulties…     

Breeding and molecular cytogenetic identification of wheat-Thinopyrum intermedium addition lines resistant to powdery mildew

Wheat-related species Th. intermedium was used to cross with common wheat Yannong 15. In the self progenies of the hybrid, two addition lines, Ⅱ-1-7-1 and Ⅱ-3-3-2, stable in cytology, were developed by cytology and powdery mildew resistance identification. Their chromosome number were 2n = 44 and formed 22 bivalents at PMC MI. In F₁ of the two addition lines crossing with Yannong 15, there appeared about one univalent at PMC MI, respectively. Resistance identification in greenhouse and field using the No. 15 and mixed strains of E. gramnis f. sp. tritici showed that they were immune to powdery mildew. Chromosome number and resistance identification using the F² single plants of the addition line crossing with Yannong 15 indicated that the resistant gene was located on the alien chromosomes. In situ hybridization using St and E genomic DNA as probe showed that the added chromosome in the two addition lines probably came from the E genome of Th. intermedium, which indicated that a pair of E genome chromosomes carried a new resistant gene to powdery mildew.     

The inhibiting effect of 1·4 recombinant P chromosome of wheat-<Emphasis Type="Italic">Agropyron cristatum</Emphasis> addition line on the <Emphasis Type="Italic">Ph</Emphasis> gene

P chromosomes may carry a genetic system that inhibits the Ph gene in wheat. Abnormal chromosome synapsis in wheat-Agropyron cristatum addition line II-21-2 (additional 1·4 recombinant P chromosome) was observed in this study. The results of cytogenetics and Ph1 gene amplification showed that the Ph1 gene was normal and the average number of quadrivalents or hexavalents was determined to be 0.41 and 0.13, respectively, in pollen-mother cells of wheat-Agropyron cristatum addition line II-21-2. The analysis o...     

小麦中与低脱氧雪腐镰刀菌烯醇含量关联的抗性基因类似物

脱氧雪腐镰刀菌烯醇(Deoxynivalenol,DON)是小麦遭受赤霉病危害后由赤霉病菌———禾谷镰刀菌产生的一种真菌毒素,这种毒素对人类和家畜的健康产生危害,严重地影响小麦的利用价值.种植低DON含量的品种是减少这种毒素危害的有效措施,而分子标记辅助育种技术能加速低DON品种的选育进程.在采用AFLP、SSR分子标记的基础上,进一步利用抗性基因类似物(RGA)对‘宁7840’(低DON含量的品种)和‘Clark’(高DON含量的品种)建立的133个重组自交系进行了DON含量的遗传特性分析.研究结果发现两个DON含量的数量特性位点(QTL),它们中含有4个与低DON含量密切相关RGAs,其中三个RGAs(RGA14-1,RGA16-1,RGA18-1)座落在染色体1A的长臂上,另一个(RGA14-2)座落在染色体5B的长臂上.座落在1AL和5BL上的QTL分别解释16.81%和7.84%的DON含量变异.DNA序列分析表明这些RGAs均与现已发现的抗性基因同源/类似,它们可能在小麦的低DON含量和赤霉病抗性上扮演着重要的角色.     

利用分子标记研究小麦遗传群体的赤霉病抗性鉴定方法

针对小麦赤霉病抗源苏麦3号构建的两个小麦重组自交系遗传群体苏麦3号／Alondra和苏麦3号／安农8455，采用单花接种、表土接种及自然发病3种不同的接种方法进行小麦赤霉病抗性接种鉴定，并根据苏麦3号赤霉病抗性主效QTL的连锁分子标记Xgwm 493和Xgwm 533．1分别对群体进行抗性连锁分析．检测结果表明，在温室单花接种所获得的鉴定数据中，标记的赤霉病抗性连锁效应最高，P值分别小于0．0001，抗性鉴定结果最为准确．研究表明，对小麦赤霉病这种由数量性状控制，受外界环境影响较大的真菌病害进行抗扩展性的遗传研究，应采用控温控湿条件下的单花滴注接种鉴定方法最为合适．     

基于抗体的小麦赤霉病抗性研究进展

 由镰刀菌引起的小麦赤霉病,不仅降低产量和品质,所形成的真菌毒素还会影响食品安全。植物缺乏高抗小麦赤霉病的资源,因此从动物中筛选、分离抗病抗体及其基因,可为培育植物抗病品种提供新种质,也拓展了开发研究新抗源的渠道。抗病抗体能特异靶向真菌蛋白、抑制抗原的功能;抗体与抗菌肽和顺式表达调控元件结合,使病菌能诱导抗体融合蛋白在镰刀菌侵染小麦的颖壳高效表达,从而有效抑制病菌侵入、扩展及毒素积累。本文综述抗病抗体的抗原类型、抗病抗体及其融合蛋白在体外和植物中的表达以功能、抗原基因结构与功能、抗体抗病机理的研究进展。     

小麦赤霉病:从表型鉴定到抗性改良

 赤霉病已成为影响中国小麦安全生产的最重要病害之一。表型鉴定体系对赤霉病抗性准确评价和抗性改良至关重要。本文分析了赤霉病的危害和改良的迫切性、中国抗赤育种的现状、QTL的定位和利用进展等,讨论了赤霉病接种鉴定和评价方法、赤霉病抗源利用和抗赤分子标记辅助育种策略,提出赤霉病改良的思路和建议。     

小麦液泡质子焦磷酸酶基因EST的克隆与分析

通过RT-PCR的方法在矮苏3小麦的总cDNA中克隆到一个与大麦液泡质子焦磷酸酶基因(VP)高度同源的EST。以该序列为基础,利用生物信息学的方法构建了一个编码小麦VP蛋白的全长EST重叠群,长2764bp,其包含一个长2355bp的完整开放读码框(ORF),编码785个氨基酸多肽。通过Southern杂交,将该VP基因定位在小麦染色体的第7同源群上。     

Breeding and molecular cytogenetic identification of wheat-Thinopyrum intermedium addition lines resistant to powdery mildew

Wheat-related species Th. intermedium was used to cross with common wheat Yannong 15. In the self progenies of the hybrid, two addition lines, Ⅱ-1-7-1 and Ⅱ-3-3-2, stable in cytology, were developed by cytology and powdery mildew resistance identification. Their chromosome number were 2n = 44 and formed 22 bivalents at PMC MI. In F1 of the two addition lines crossing with Yannong 15, there appeared about one univalent at PMC MI, respectively. Resistance identification in greenhouse and field using the No. 15 and mixed strains of E. gramnis f. sp. tritici showed that they were immune to powdery mildew. Chromosome number and resistance identification using the F2 single plants of the addition line crossing with Yannong 15 indicated that the resistant gene was located on the alien chromosomes. In situ hybridization using St and E genomic DNA as probe showed that the added chromosome in the two addition lines probably came from the E genome of Th. intermedium, which indicated that a pair of E genome chromosomes carried a new resistant gene to powdery mildew.     

利用小麦5B效应引入外源基因的研究（Ⅰ）易位系的培育

ｄｕｒｕｍ小麦的代换系ｄｉ－ｓｕｂ５Ｄ（５Ｂ）与添加系ｄｉ－ａｄｄｅ４ｔｓ杂交，再用ｄｉ－ｓｕｂ５Ｄ（５Ｂ）进行回交，在自交后代中选育出了易位系１０３２。该易位系染色体数２ｎ＝２８，表现型为非蜡质。这一结果证明了在ｄｕｒｕｍ小麦中也可以利用５Ｂ染色体效应，通过诱发部份同源染色体间的配对，获得易位体。