Weihua Liu

The introgression of chromosome 6P specifying for increased numbers of florets and kernels from Agropyron cristatum into wheat.

A wheat (Triticum aestivum L.) line 4844 with superior numbers of florets and grains per spike was derived from the cross between Fukohokomugi wheat and Agropyron cristatum (L.) Gaertn. In order to determine the genetic control of floret and kernel number per spike in this line, chromosome addition and substitution lines that were derived from line 4844 were characterized by means of in situ hybridization, microsatellite (SSR), and gliadin analyses. Genomic in situ hybridization analysis with biotinylated P genomic DNA of A. cristatum as a probe demonstrated that the increased number of florets and grains in a spike was associated with the introgression of an A. cristatum chromosome. Fluorescence in situ hybridization, using a repetitive sequence, pAs1, derived from Aegilops squarrosa L., indicated the replacement of chromosome 6D of wheat in the wheat-A. cristatum chromosome substitution lines. This was confirmed by microsatellite analyses with wheat SSR markers specific for chromosome 6D, suggesting that the A. cristatum chromosome was homoeologous to group 6 and was therefore designated as 6P. This conclvsion was further confirmed by amplification using EST-SSR markers and gliadin analysis. The increased number of florets and kernels within a spike of the wheat-A. cristatum hybrids thus was controlled by gene(s) located on A. cristatum chromosome 6P.

QTL mapping of yield-related traits in the wheat germplasm 3228

The new wheat germplasm 3228, a putative derivative of tetraploid Agropyron cristatum Z559 and the common wheat Fukuhokomugi, has superior features in yield-related traits, particularly in spike morphological traits, such as large spike and superior grain number. To identify favorable alleles of these traits in 3228, 237 F2:3 families were developed from the cross 3228/Jing 4839. A genetic map was constructed using 179 polymorphic SSR and EST-SSR markers. A total of 76 QTL controlling spike number per plant (SNP), spike length (SL), spikelet number per spike (SNS), floret number per spikelet (FNS), grain number per spike (GNS) and thousand-grain weight (TGW) were detected on 16 chromosomes. Each QTL explained 1.24–27.01% of the phenotypic variation, and 9 QTL (28.95%) were detected in two or all environments. Additive effects of 45 QTL were positive with 3228 alleles increasing the QTL effects, 31 QTL had negative effects indicating positive contributions from Jing 4839. Three important clusters involving all traits were located on chromosomes 5A, 6A and 4B, and several co-located QTL were also found. Most of the QTL detected on the three chromosome regions could contribute to the use of 3228 in breeding for grain yield improvement.

Cloning and characterization of repetitive sequences and development of SCAR markers specific for the P genome of Agropyron cristatum

In previous studies, we successfully transferred the P genome of Agropyron cristatum into wheat using wide hybridization methods. In the current investigation, repetitive sequences were cloned and DNA markers specific for the P genome of A. cristatum were developed. Three P genome-specific markers, designated OPX07-1036, OPX11-817 and OPC05-1539, were identified and isolated using random amplified polymorphic DNAs. The three markers were successfully amplified in all tested materials that contained Agropyron chromatin, such as Agropyron itself and wheat-Agropyron addition lines. These RAPD markers were converted into SCAR markers to be used in detection of P genome chromatin in wheat. In situ probing with fluorescent-labeled marker DNA has shown that they are distributed in all arms of Agropyron hence they will be useful in a variety of studies on introgressions of the P-genome chromatin into wheat.

Introgression of Agropyron cristatum 6P chromosome segment into common wheat for enhanced thousand-grain weight and spike length

Key messageThis study explored the genetic constitutions of wheat-Agropyron cristatum6P chromosomal translocation and determined the effects of 6P intercalary chromosome segment on thousand-grain weight and spike length in wheat.AbstractCrop wild relatives provide rich genetic resources for wheat improvement. Introduction of alien genes from Agropyron cristatum into common wheat can broaden its genetic diversity. In this study, radiation-induced wheat-A. cristatum translocation line Pubing3035 derived from the offspring of wheat-A. cristatum 6P chromosomes addition line was identified and analyzed using genomic in situ hybridization (GISH), dual-color fluorescence in situ hybridization (FISH), and molecular markers. GISH analysis revealed that Pubing3035 was a Ti1AS-6PL-1AS·1AL intercalary translocation. The breakpoint was pinpointed to locate near the centromeric region on the short arm of wheat chromosome 1A based on a constructed F2 linkage map and it was flanked by markers SSR12 and SSR263. The genotypic data, combined with the phenotypes, indicated that A. cristatum 6P chromosomal segment played an important role in regulating the thousand-grain weight and spike length. On average, the thousand-grain weight and spike length in translocation individuals were approximately 2.5xa0g higher and 0.7xa0cm longer than those in non-translocation individuals in F2 and BC1F1 populations. The clusters of quantitative trait loci for thousand-grain weight, spike length, and spikelet density contributed by 6P chromosome segment were mapped between A. cristatum unique marker Agc7155 and wheat marker SSR263, which, respectively, explained phenotypic variance of 24.96, 12.38 and 17.20xa0% with an LOD of 10.63, 4.89 and 5.59. Overall, the translocation Pubing3035 had a positive effect on the yield of wheat, which laid the foundation for the localization of A. cristatum excellent genes and made itself a promising and valuable germplasm for wheat improvement.

The effects of chromosome 6P on fertile tiller number of wheat as revealed in wheat-Agropyron cristatum chromosome 5A/6P translocation lines

Key messageThis study explored the genetic constitutions of several wheat-A. cristatum translocation lines and determined the effects of A. cristatum6P chromosome segments on fertile tiller number in wheat.AbstractProgress in wheat breeding is hampered by a relatively narrow range of genetic variation. To overcome this hurdle, wild relatives of common wheat with superior agronomic traits are often used as donors of desirable genes in wheat-breeding programs. One of the successfully utilized wheat wild relatives is Agropyron cristatum (L.) Gaertn (2nxa0=xa04xxa0=xa028; genomes PPPP). We previously reported that WAT31-13 was a wheat-A. cristatum 5A-6P reciprocal translocation line with higher fertile tiller number and grain number per spike compared to common wheat. However, WAT31-13 was genetically unstable. In this study, we analyzed the 43 genetically stable progenies from WAT31-13 using genomic in situ hybridization, dual-color fluorescence in situ hybridization, and molecular markers. We classified them into three translocation types (TrS, TrL and TrA) and seven subtypes, and also pinpointed the translocation breakpoint. The genotypic data, combined with the phenotypes of each translocation type, enabled us to physically map agronomic traits to specific A. cristatum 6P chromosome arms or segments. Our results indicated that A. cristatum chromosome 6P played an important role in regulating fertile tiller number, and that positive and negative regulators of fertile tiller number existed on the A. cristatum chromosome arm 6PS and 6PL, respectively. By exploring the relationship between fertile tiller number and A. cristatum chromosome segment, this study presented a number of feasible approaches for creation, analysis, and utilization of wheat-alien chromosome translocation lines in genetic improvement of wheat.

Physical mapping of Agropyron cristatum chromosome 6P using deletion lines in common wheat background

Key messageGenetically stable deletion lines ofAgropyron cristatumchromosome 6P in common wheat background were generated, which allowed for physical mapping of 255 6P-specific STS markers and leaf rust resistance gene(s).AbstractChromosomal deletion lines are valuable tools for gene discovery and localization. The chromosome 6P of Agropyron cristatum (2nxa0=xa04xxa0=xa028, PPPP) confers many desirable agronomic traits to common wheat, such as higher grain number per spike, multiple fertile tiller number, and enhanced resistance to certain diseases. Although many elite genes from A. cristatum have been identified, their chromosomal locations were largely undetermined due to the lack of A. cristatum 6P deletion lines. In this study, various A. cristatum 6P deletion lines were developed using a wheat–A. cristatum 6P disomic addition line 4844-12 subjected to 60Co-γ irradiation as well as an Aegilops cylindrica gametocidal chromosome. Twenty-six genetically stable A. cristatum 6P deletion lines in the genetic background of common wheat were obtained, and their genetic constitutions were elucidated by genomic in situ hybridization (GISH) and sequence-tagged site (STS) markers specific to A. cristatum chromosome 6P. Moreover, 255 novel chromosome 6P-specific STS markers were physically mapped to 14 regions of chromosome 6P. Field evaluation of leaf rust resistance of various deletion lines and BC1F2 populations indicated that the A.cristatum chromosome 6P-originated leaf rust resistance gene(s) was located in the region 6PS-0.81-1.00. This study will provide not only useful tools for characterization and utilization of wheat materials with alien chromosomal segments, but also novel wheat germplasms potentially valuable in wheat breeding and improvement.

Mapping of novel powdery mildew resistance gene(s) from Agropyron cristatum chromosome 2P.

Key messageA physical map ofAgropyron cristatum2P chromosome was constructed for the first time and the novel powdery mildew resistance gene(s) from chromosome 2P was(were) also mapped.AbstractAgropyron cristatum (L.) Gaertn. (2nxa0=xa028, PPPP), a wild relative of common wheat, is highly resistant to powdery mildew. Previous studies showed that wheat-A. cristatum 2P disomic addition line II-9-3 displayed high resistance to powdery mildew, and the resistance was attributable to A. cristatum chromosome 2P. To utilize and physically map the powdery mildew resistance gene(s), 15 wheat-A. cristatum 2P translocation lines and three A. cristatum 2P deletion lines with different chromosomal segment sizes, obtained from II-9-3 using 60Co-γ ray irradiation, were characterized using cytogenetic and molecular marker analysis. A. cristatum 2P chromosomal segments in the translocations were translocated to different wheat chromosomes, including 1A, 4A, 5A, 6A, 7A, 1B, 2B, 3B, 7B, 3D, 4D, and 6D. A physical map of the 2P chromosome was constructed with 82 STS markers, consisting of nine bins with 34 markers on 2PS and eight bins with 48 markers on 2PL. The BC1F2 populations of seven wheat-A. cristatum 2P translocation lines (2PT-3, 2PT-4, 2PT-5, 2PT-6, 2PT-8, 2PT-9, and 2PT-10) were developed by self-pollination, tested with powdery mildew and genotyped with 2P-specific STS markers. From these results, the gene(s) conferring powdery mildew resistance was(were) located on 2PL bin FL 0.66–0.86 and 19 2P-specific markers were identified in this bin. Moreover, two new powdery mildew-resistant translocation lines (2PT-4 and 2PT-5) with small 2PL chromosome segments were obtained. The newly developed wheat lines with powdery mildew resistance and the closely linked molecular markers will be valuable for wheat disease breeding in the future.

Molecular cytogenetic characterization of an Agropyron cristatum 6PL chromosome segment conferring superior kernel traits in wheat

The wild relative of wheat, Agropyron cristatum (L.) Gaertn. (genome PPPP), acts as an important genetic resource for providing valuable genes for wheat improvement and increasing the diversity of cultivated wheat. In the present study, cytogenetic and molecular tools were used to characterize wheat-A. cristatum 6P derivatives. The novel translocation line Pubing260 exhibiting wider flag leaves and superior spike traits was further characterized and analysed. Cytological studies demonstrated that Pubing260 (2nu2009=u200942) contained a T3BL·3BS-6PL terminal translocation. We compared molecular markers on chromosomes 6P and 3B and confirmed that the translocated wheat chromosome was 3BS and that the chromosome breakage occurred in bin 3BS9-0.57-0.75. Compared with its recurrent parent Fukuhokomugi, Pubing260 had a wider flag leaf, more spikelets and more grains per spike in two growing seasons. Genetic analysis conducted using BC1F2 and BC2F1 populations suggested that the A. cristatum chromosomal fragment from bin 6PL-0.72-1.00 conferred these potentially valuable agronomic traits. On average, the flag leaf width (FLW), and numbers of grain per spike (GNS), spikelets per spike (SNS) and kernels per spikelet of plants with the translocation were 2xa0mm wider and 5.1, 0.8 and 0.3 higher, respectively, than those of plants lacking the translocation in segregating populations. Significant and positive correlations were observed among GNS, SNS and FLW. In summary, this study not only reports a novel germplasm that is potentially valuable for wheat improvement but also enriches the genetic resources of wheat.