Cytogenetic and Molecular Marker Analyses of a Novel Wheat-Psathyrostachys huashanica 7Ns Disomic Addition Line with Powdery Mildew Resistance.

Powdery mildew caused by Blumeria graminis f. sp. tritici is a devastating disease that reduces wheat yield and quality worldwide. The exploration and utilization of new resistance genes from wild wheat relatives is the most effective strategy against this disease. Psathyrostachys huashanica Keng f. ex P. C. Kuo (2n = 2x = 14, NsNs) is an important tertiary gene donor with multiple valuable traits for wheat genetic improvement, especially disease resistance. In this study, we developed and identified a new wheat-P. huashanica disomic addition line, 18-1-5-derived from a cross between P. huashanica and common wheat lines Chinese Spring and CSph2b. Sequential genomic and multicolor fluorescence in situ hybridization analyses revealed that 18-1-5 harbored 21 pairs of wheat chromosomes plus a pair of alien Ns chromosomes. Non-denaturing fluorescence in situ hybridization and molecular marker analyses further demonstrated that the alien chromosomes were derived from chromosome 7Ns of P. huashanica. The assessment of powdery mildew response revealed that line 18-1-5 was highly resistant at the adult stage to powdery mildew pathogens prevalent in China. The evaluation of agronomic traits indicated that 18-1-5 had a significantly reduced plant height and an increased kernel length compared with its wheat parents. Using genotyping-by-sequencing technology, we developed 118 PCR-based markers specifically for chromosome 7Ns of P. huashanica and found that 26 of these markers could be used to distinguish the genomes of P. huashanica and other wheat-related species. Line 18-1-5 can therefore serve as a promising bridging parent for wheat disease resistance breeding. These markers should be conducive for the rapid, precise detection of P. huashanica chromosomes and chromosomal segments carrying Pm resistance gene(s) during marker-assisted breeding and for the investigation of genetic differences and phylogenetic relationships among diverse Ns genomes and other closely related ones.

Genetic diversity of Asian and European common wheat lines assessed by fluorescence in situ hybridization.

Understanding the genetic diversity of wheat is important for wheat breeding and improvement. However, there have been limited attempts to evaluate wheat diversity using fluorescence in situ hybridization (FISH). In this study, the chromosomal structures of 149 wheat accessions from 13 countries located between the latitudes of 30°N and 45°N, the principal growing region for wheat, were characterized using FISH with pTa535 and pSc119.2 probes. The ranges of the numbers of FISH types in the A-, B-, and D-genome chromosomes were 2-8, 3-7, and 2-4, respectively, and the average numbers in the A and B genomes were greater than in the D genome. Chromosomal translocations were detected by these probes, and previously undescribed translocations were also observed. Using the FISH, the genetic relationships among the 149 common wheat lines were divided into three groups (G1, G2, and G3). G1 mainly consisted of southern European lines, G2 consisted of most lines from Japan and some lines from western Asia, China, and Korea, and G3 consisted of the other lines from southern Europe and most of the lines from western Asia, China, and Korea. FISH karyotypes of wheat chromosomes distinguished chromosomal structural variations, revealing the genetic diversity among wheat varieties. Furthermore, these results provide valuable information for the further genetic improvement of wheat in China.

Characterization of a wheat-tetraploid Thinopyrum elongatum 1E(1D) substitution line K17-841-1 by cytological and phenotypic analysis and developed molecular markers.

BACKGROUND: Tetraploid Thinopyrum elongatum (2n = 4x = 28) is a promising source of useful genes, including those related to adaptability and resistance to diverse biotic (Fusarium head blight, rust, powdery mildew, and yellow dwarf virus) and abiotic (cold, drought, and salt) stresses. However, gene transfer rates are low for this species and relatively few species-specific molecular markers are available. RESULTS: The wheat-tetraploid Th. elongatum line K17-841-1 derived from a cross between a hexaploid Trititrigia and Sichuan wheat cultivars was characterized based on sequential genomic and fluorescence in situ hybridizations and simple sequence repeat markers. We revealed that K17-841-1 is a 1E (1D) chromosomal substitution line that is highly resistant to stripe rust pathogen strains prevalent in China. By comparing the sequences generated during genotyping-by-sequencing (GBS), we obtained 597 specific fragments on the 1E chromosome of tetraploid Th. elongatum. A total of 235 primers were designed and 165 new Th. elongatum-specific markers were developed, with an efficiency of up to 70%. Marker validation analyses indicated that 25 specific markers can discriminate between the tetraploid Th. elongatum chromosomes and the chromosomes of other wheat-related species. An evaluation of the utility of these markers in a F2 breeding population suggested these markers are linked to the stripe rust resistance gene on chromosome 1E. Furthermore, 28 markers are unique to diploid Th. elongatum, tetraploid Th. elongatum, or decaploid Thinopyrum ponticum, which carry the E genome. Finally, 48 and 74 markers revealed polymorphisms between Thinopyrum E-genome- containing species and Thinopyrum bessarabicum (Eb) and Pseudoroegneria libanotica (St), respectively. CONCLUSIONS: This new substitution line provide appropriate bridge-breeding-materials for alien gene introgression to improve wheat stripe rust resistance. The markers developed using GBS technology in this study may be useful for the high-throughput and accurate detection of tetraploid Th. elongatum DNA in diverse materials. They may also be relevant for investigating the genetic differences and phylogenetic relationships among E, Eb, St, and other closely-related genomes and for further characterizing these complex species.

Identification of a Wheat-Psathyrostachys huashanica 7Ns Ditelosomic Addition Line Conferring Early Maturation by Cytological Analysis and Newly Developed Molecular and FISH Markers.

Early maturation is an important objective in wheat breeding programs that could facilitate multiple-cropping systems, decrease disaster- and disease-related losses, ensure stable wheat production, and increase economic benefits. Exploitation of novel germplasm from wild relatives of wheat is an effective means of breeding for early maturity. Psathyrostachys huashanica Keng f. ex P. C. KUO (2n=2x=14, NsNs) is a promising source of useful genes for wheat genetic improvement. In this study, we characterized a novel wheat-P. huashanica line, DT23, derived from distant hybridization between common wheat and P. huashanica. Fluorescence in situ hybridization (FISH) and sequential genomic in situ hybridization (GISH) analyses indicated that DT23 is a stable wheat-P. huashanica ditelosomic addition line. FISH painting and PCR-based landmark unique gene markers analyses further revealed that DT23 is a wheat-P. huashanica 7Ns ditelosomic addition line. Observation of spike differentiation and the growth period revealed that DT23 exhibited earlier maturation than the wheat parents. This is the first report of new earliness per se (Eps) gene(s) probably associated with a group 7 chromosome of P. huashanica. Based on specific locus-amplified fragment sequencing technology, 45 new specific molecular markers and 19 specific FISH probes were developed for the P. huashanica 7Ns chromosome. Marker validation analyses revealed that two specific markers distinguished the Ns genome chromosomes of P. huashanica and the chromosomes of other wheat-related species. These newly developed FISH probes specifically detected Ns genome chromosomes of P. huashanica in the wheat background. The DT23 line will be useful for breeding early maturing wheat. The specific markers and FISH probes developed in this study can be used to detect and trace P. huashanica chromosomes and chromosomal segments carrying elite genes in diverse materials.

Genetic Diversity and Population Structure of Asian and European Common Wheat Accessions Based on Genotyping-By-Sequencing.

Obtaining information on the genetic diversity and population structure of germplasm facilitates its use in wheat breeding programs. Recently, with the development of next-generation sequencing technology, genotyping-by-sequencing (GBS) has been used as a high-throughput and cost-effective molecular tool for examination of the genetic diversity of wheat breeding lines. In this study, GBS was used to characterize a population of 180 accessions of common wheat originating from Asia and Europe between the latitudes 30° and 45°N. In total, 24,767 high-quality single-nucleotide polymorphism (SNP) markers were used for analysis of genetic diversity and population structure. The B genome contained the highest number of SNPs, followed by the A and D genomes. The polymorphism information content was in the range of 0.1 to 0.4, with a mean of 0.26. The distribution of SNPs markers on the 21 chromosomes ranged from 243 on chromosome 4D to 2,337 on chromosome 3B. Structure and cluster analyses divided the panel of accessions into two subgroups (G1 and G2). G1 principally consisted of European and partial Asian accessions, and G2 comprised mainly accessions from the Middle East and partial Asia. Molecular analysis of variance showed that the genetic variation was greater within groups (99%) than between groups (1%). Comparison of the two subgroups indicated that G1 and G2 contained a high level of genetic diversity. The genetic diversity of G2 was slightly higher as indicated by the observed heterozygosity (H o) = 0.23, and unbiased diversity index (uh) = 0.34. The present results will not only help breeders to understand the genetic diversity of wheat germplasm on the Eurasian continent between the latitudes of 30° and 45°N, but also provide valuable information for wheat genetic improvement through introgression of novel genetic variation in this region.