Discrimination of Repetitive Sequences Polymorphism in Secale cereale by Genomic In Situ Hybridization-Banding

Genomic in situ hybridization banding （GISH-banding）, a technique slightly modified from conventional GISH, was used to probe the Chinese native rye （Secale cereale L.） DNA, and enabled us to visualize the individual rye chromosomes and create a universal reference karyotype of the S. cereale chromosome 1R to 7R. The GISH-banding approach used in the present study was able to discriminate S. cereale chromosomes or segments in the wheat （Triticum aestivum L.） background, including the Triticale, wheat-rye addition and translocation lines. Moreover, the GISH-banding pattern of S. cereale subsp. Afghanicum chromosomes was consistent with that of Chinese native rye cv. Jingzhou rye; whereas the GISH-banding pattern of Secale vavilovii was different from that of S. cereale, indicating that GISH-banding can be used to study evolutionary polymorphism in species or subspecies of Secale. In addition, the production and application of GISH-banding to the study of adenine-thymine-riched heterochromaUn is discussed.

Development and identification of two novel wheat-rye 6R derivative lines with adult-plant resistance to powdery mildew and high-yielding potential

Powdery mildew,caused by Blumeria graminis f.sp.tritici(Bgt),is a devastating disease that seriously threatens wheat yield and quality.To control this disease,host resistance is the most effective measure.Compared with the resistance genes from common wheat,alien resistance genes can better withstand infection of this highly variable pathogen.Development of elite alien germplasm resources with powdery mildew resistance and other key breeding traits is an attractive strategy in wheat breeding.In this study,three wheat-rye germplasm lines YT4-1,YT4-2,and YT4-3 were developed through hybridization between octoploid triticale and common wheat,out of which the lines YT4-1 and YT4-2 conferred adult-plant resistance(APR)to powdery mildew while the line YT4-3 was susceptible to powdery mildew during all of its growth stages.Using genomic in situ hybridization,multi-color fluorescence in situ hybridization,multi-color GISH,and molecular marker analysis,YT4-1,YT4-2,and YT4-3 were shown to be cytogenetically stable wheat-rye 6R addition and T1RS.1BL translocation line,6RL ditelosomic addition and T1RS.1BL translocation line,and T1RS.1BL translocation line,respectively.Compared with previously reported wheat-rye derivative lines carrying chromosome 6R,YT4-1 and YT4-2 showed stable APR without undesirable pleiotropic effects on agronomic traits.Therefore,these novel wheat-rye 6R derivative lines are expected to be promising bridge resources in wheat disease breeding.

Molecular cytogenetic analyses of two new wheat-rye 6RL translocation lines with resistance to wheat powdery mildew

Rye(Secale cereale)is a valuable gene donor for wheat improvement,especially for its resistance to diseases.Developing rye-derived resistance sources is important for wheat breeding.In the present study,two wheat-rye derivatives,designated JS016 and JS110,were produced by crossing common wheat cultivar Yangmai 23 with Pakistani rye accession W2A.Using sequential genomic in situ hybridization(GISH)and multicolor fluorescence in situ hybridization(mc-FISH),JS016 and JS110 were identified as a T6BS.6RL translocation line and a T6BS.6BL6RL translocation line,respectively.Ten newly 6RL chromosome arm-specific markers were developed and used to confirm the 6RL translocation.The wheat 55K single-nucleotide polymorphism(SNP)array further verified the molecular cytogenetic identification results above and clarified their breakpoints at 430.9 and 523.0 Mb of chromosome 6B in JS016 and JS110,respectively.Resistance spectrum and allelism test demonstrated that JS016 and JS110 possessed novel powdery mildew resistance gene(s)that was derived from the 6RL translocation but differed from Pm20.Moreover,JS016 and JS110 had better agronomic traits than the previously reported 6RL translocation line carrying Pm20.To efficiently transfer and detect the 6RL translocation from JS016 and JS110,one 6RL-specific Kompetitive allele specific PCR(KASP)marker was developed and validated in high throughput marker-assisted selection(MAS).

Cytological and genetic analyses of a wheat-rye 2RL ditelosomic addition line with adult plant resistance to powdery mildew

Rye(Secale cereale genome RR),a close relative of common wheat,possesses valuable resistance genes for wheat improvement.Due to the co-evolution of pathogen virulence and host resistance,some resistance genes derived from rye have lost effectiveness.Development and identification of new,effective resistance genes from rye is thus required.In the current study,wheat-rye line WR56 was produced through distant hybridization,embryo rescue culture,chromosome doubling and backcrossing.WR56 was then proved to be a wheat-rye 2 RL ditelosomic addition line using GISH(genomic in situ hybridization),mc-FISH(multicolor fluorescence in situ hybridization),ND-FISH(non-denaturing FISH),mc-GISH(multicolor GISH)and rye chromosome arm-specific marker analysis.WR56 exhibited a high level of adult plant resistance to powdery mildew caused by Blumeria graminis f.sp.tritici(Bgt).This resistance was carried by the added 2 RL telosomes and presumed to be different from Pm7 which is also located on chromosome arm 2 RL but confers resistance at the seedling and adult stages.WR56 will be a promising bridging parent for transfer of the resistance to a more stable wheat breeding line.A newly developed2 RL-specific KASP(kompetitive allele specific PCR)marker should expedite that work.

Identification of genes involved in the formation of soluble dietary fiber in winter rye grain and their expression in cultivars with different viscosities of wholemeal water extract

The grain of rye(Secale cereale L.) used for baking contains a large amount of non-starch polysaccharides,making it an excellent component of functional foods. But rye grain intended for alcohol production and forage use should have a reduced content of these polysaccharides. A comprehensive parameter that can predict the best field of application for winter rye grain is the viscosity of its wholemeal water extract.However, our understanding of the genetic background underlying this key trait and associated features of rye grain is poor. By analyzing six Russian winter rye cultivars, we identified the most contrasting forms and characterized the peculiarities of their water-soluble carbohydrates capable of influencing the viscosity of water extracts. Then, using phylogenetic and transcriptomic analyses, we identified in the rye genome many genes encoding putative glycosyltransferases and glycosylhydrolases responsible for the synthesis and degradation of arabinoxylans, mixed-linkage glucans, cellulose, and some other polysaccharides. We determined the dynamics of m RNA abundance for these genes at three stages of kernel development. Comparisons of gene expression levels in two contrasting cultivars revealed specific members of multigene families that may serve as promising targets for manipulating non-starch polysaccharide content in rye grain. High-viscosity cultivars were characterized by up-regulation of many glycosyltransferases involved in the biosynthesis of arabinoxylans and other cell-wall polysaccharides,whereas low-viscosity cultivars showed up-regulation of several genes encoding polysaccharidedegrading enzymes.

三种黑麦种质的染色体FISH核型分析

为了解黑麦染色体的FISH核型特点,本研究使用Oligo-pSc119.2-2、Oligo-pSc200和Oligo-pSc250探针对3种黑麦(MAD黑麦, WR7黑麦和C718黑麦)染色体进行了双色FISH分析,结果发现3份黑麦的7对染色体上,Oligo-pSc119.2-2红色信号强且丰富,广泛分布于染色体端部及中部等。Oligo-pSc200和Oligo-pSc250绿色信号强,主要位于染色体端部及近端部。3份黑麦每条染色体的Oligo-pSc119.2-2信号分布较为相似,但是Oligo-pSc200和Oligo-pSc250信号的差别较大。MAD黑麦和WR7黑麦染色体上的Oligo-pSc200和Oligo-pSc250信号比C718黑麦多且强。MAD黑麦和WR7黑麦的FISH核型较为相似,它们与C718黑麦的差别较大。通过本试验了解到3份黑麦的FISH核型之间存在遗传多样性,建立了3份黑麦的FISH核型,这将有助于该种质在麦类作物遗传育种上的利用和深入研究。

Population genomic analysis reveals domestication of cultivated rye from weedy rye

Rye(Secale cereale)is an important crop with multiple uses and a valuable genetic resource for wheat breeding.However,due to its complex genome and outcrossing nature,the origin of cultivated rye remains elusive.The geneticist N.I.Vavilov proposed that cultivated rye had been domesticated from weedy rye,rather than directly from wild species like other crops.Unraveling the domestication history of rye will extend our understanding of crop evolution and upend our inherent understanding of agricultural weeds.To this end,in this study we generated the 8.5 Tb of whole-genome resequencing data from 116 worldwide accessions of wild,weedy,and cultivated rye,and demonstrated that cultivated rye was domesticated directly from weedy relatives with a similar but enhanced genomic selection by humans.We found that a repertoire of genes that experienced artificial selection is associated with important agronomic traits,including shattering,grain yield,and disease resistance.Furthermore,we identified a composite introgression in cultivated rye from the wild perennial Secale strictum and detected a 2-Mb introgressed fragment containing a candidate ammonium transporter gene with potential effect on the grain yield and plant growth of rye.Taken together,our findings unravel the domestication history of cultivated rye,suggest that interspecific introgression serves as one of the likely causes of obscure species taxonomy of the genus Secale,and provide an important resource for future rye and wheat breeding.

Variation of B Chromosome Associated with Tissue Culture in Wheat-rye Cross

In vitro variation of B chromosomes was studied by examining the callus cells derived from the immature embryos from a cross of Chinese Spring wheat （Triticum aestivum L.） and Fin 7416 rye （Secale cereale L.） carrying two B chromosomes. In 40-d-old callus cells, the numbers of B chromosomes ranged from one to four in 65.6% of the cells observed. The distribution of B chromosome numbers was associated with the ploidy levels of the normal chromosomes （A chromosomes）. The frequency of the cells with high numbers of B chromosomes （i.e., three or four B chromosomes） in the amphiploid cells with 56 A chromosomes was greater than those in the haploid cells with 28 A chromosomes. Although structural changes in the rye A chromosomes were observed, cytological observation and genomic in situ hybridization demonstrated that the rye B chromosomes were conserved in morphological appearance following tissue culture.