Nobuyoshi Watanabe

FRIZZY PANICLE Drives Supernumerary Spikelets in Bread Wheat

Wheat transcription factors located on chromosome group 2 drive the yield-related production of supernumerary spikelets. Bread wheat (Triticum aestivum) inflorescences, or spikes, are characteristically unbranched and normally bear one spikelet per rachis node. Wheat mutants on which supernumerary spikelets (SSs) develop are particularly useful resources for work towards understanding the genetic mechanisms underlying wheat inflorescence architecture and, ultimately, yield components. Here, we report the characterization of genetically unrelated mutants leading to the identification of the wheat FRIZZY PANICLE (FZP) gene, encoding a member of the APETALA2/Ethylene Response Factor transcription factor family, which drives the SS trait in bread wheat. Structural and functional characterization of the three wheat FZP homoeologous genes (WFZP) revealed that coding mutations of WFZP-D cause the SS phenotype, with the most severe effect when WFZP-D lesions are combined with a frameshift mutation in WFZP-A. We provide WFZP-based resources that may be useful for genetic manipulations with the aim of improving bread wheat yield by increasing grain number.

Genetic mapping of a mutant gene producing three pistils per floret in common wheat

A common wheat (Triticum aestivum L.) mutation that produces 3 pistils (TP) per floret may result in formation of up to 3 kernels per floret. The TP trait may be important for increasing the number of grains per spike and for improving the yield potential through breeding. This trait is determined by the dominantPis1 gene. Genetic mapping ofPis1 involved 234 microsatellite markers and bulk segregant analysis of a cross of the TP line with Novosibirskaya 67. ThePis1 gene is located on chromosome 2DL, between markersXgwm539 andXgwm349. This result does not agree with a previously published localization of thePis1 gene on chromosome 5B. The possible importance of TP wheat as an alternative genetic resource is discussed.

Genetic mapping of the gene affecting polyphenol oxidase activity in tetraploid durum wheat.

The quality of durum wheat (Triticum turgidum ssp.durum) is influenced by polyphenol oxidase (PPO) activity and its corresponding substrates. A saturated molecular-marker linkage map was constructed previously by using a set of recombinant inbred (RI) lines, derived from a cross between durum wheat cultivars Jennah Khetifa and Cham 1. Quantitative trait loci (QTL) for PPO activity in seeds were mapped in this population. PPO activity in seeds of the parents and 110 RI lines was measured spectrophotometrically. The PPO activity of Cham 1 was significantly lower than that of Jennah Khetifa. QTL analysis of these data indicated that most of PPO activity was associated with major loci on the long arm of chromosome 2A. The trait was found to be strongly associated with the SSR markerXgwm312@2A. With this knowledge, marker-assisted selection can be used to select genotypes with lower PPO activity in durum wheat populations.

New alleles of the wheat domestication gene Q reveal multiple roles in growth and reproductive development

The advantages of free threshing in wheat led to the selection of the domesticated Q allele, which is now present in almost all modern wheat varieties. Q and the pre-domestication allele, q, encode an AP2 transcription factor, with the domesticated allele conferring a free-threshing character and a subcompact (i.e. partially compact) inflorescence (spike). We demonstrate that mutations in the miR172 binding site of the Q gene are sufficient to increase transcript levels via a reduction in miRNA-dependent degradation, consistent with the conclusion that a single nucleotide polymorphism in the miRNA binding site of Q relative to q was essential in defining the modern Q allele. We describe novel gain- and loss-of-function alleles of Q and use these to define new roles for this gene in spike development. Q is required for the suppression of ‘sham ramification’, and increased Q expression can lead to the formation of ectopic florets and spikelets (specialized inflorescence branches that bear florets and grains), resulting in a deviation from the canonical spike and spikelet structures of domesticated wheat. Highlighted Article: New alleles of a wheat AP2 gene, Q, define new roles for Q in inflorescence architecture, including the formation and structure of grain-producing spikelets. See also Debernardi et al. in this issue.

Q gene variability in wheat species with different spike morphology

Q gene is the major domestication gene of wheat and many questions concerning Q gene genetics, including Q gene variability and its functional influence on phenotype, remain unanswered for the majority of wheat species. Here we crossed wheat species with dominant (Q) and recessive (q) alleles and confirmed that Q gene controls threshability, rachis fragility and spike shape traits. In the present study 18 new Q gene sequences were obtained and the Q gene sequences from 42 di- and polyploid wheat species with variable spike morphology were analyzed. We identified correlation between Q gene variability (coding mutation 329Val/Ile, promoter variability, microRNA172 binding site substitution) and threshability, rachis fragility and spike shape traits in polyploid wheat species. The analysis of 3D structures of q and Q proteins indicated that 329Val/Ile mutation does not affect overall protein structure and likely protein activity. We conclude that alterations in all three regions are essential for the formation of free-threshing non fragile normal phenotype in polyploid wheat.

Exploration of genetic diversity among Xinjiang Triticum and Triticum polonicum by AFLP markers

Seventy-two XinjiangTriticum andTriticum polonicum accessions were subjected to AFLP analyses to discuss the origin ofTriticum petropavlovskyi. A total of 91 putative loci were produced by four primer combinations. Among them 56 loci were polymorphic, which is equivalent to 61.53 % of the total number of putative loci. Genetic diversity among 11T. petropavlovskyi accessions was narrow due to the lowest number (32) of polymorphic loci among the wheat species. Forty four polymorphic loci were found inT. aestivum andT. compactum, whereas the highest polymorphism was observed inT. polonicum. On the basis of the UPGMA clustering and PCO grouping and genetic similarity estimates from the AFLPs, we noted thatT. petropavlovskyi was more closely related to the Chinese accessions ofT. polonicum than toT. polonicum from other countries. Two accessions ofT. aestivum were grouped withT. petropavlovskyi in the UPGMA clustering. Both of them were similar toT. petropavlovskyi in respect of spike structure, i.e. the presence of awn, glume awn and also the presence of leaf pubescence. Six loci, which were commonly absent in ChineseT. polonicum, were also absent in almost all of theT. petropavlovskyi accessions. Findings of this study reduced the probability of an independent allopolyploidization event in the origin ofT. petropavlovskyi and indicated a greater degree of gene flow betweenT. aestivum andT. polonicum leading toT. petropavlovskyi. It is most likely that theP-gene ofT. petropavlovskyi hexaploid wheat was introduced fromT. polonicum toT. aestivum via a spontaneous introgression or breeding effort.

Comparative genetic diversity of Triticum aestivum–Triticum polonicum introgression lines with long glume and Triticum petropavlovskyi by AFLP-based assessment

AbstractsGenetic diversity of a set of introgression lines of Triticum aestivum L./T. polonicum L. with long glume and T. petropavlovskyi Udacz. et Migusch. were analyzed by Amplified Fragments Length Polymorphism (AFLP). Small-scale bulk breeding method was applied throughout until F6 generation to develop the introgression lines. Thirty-eight hexapolid F7 plants with long glume phenotype and their parents were subjected to AFLP analysis by four primer combinations. A total of 47 polymorphic loci were detected between the parents, 15 of them were introgressed across the 38 lines. It was hypothesized that approximately 50% of A or B genomes associated polymorphic loci were introgressed. The variation of introgression lines was limited within the diversity between their parents, T. aestivum L. cv. Novosibirskaya 67 (N67) and T. polonicum L. cv. IC12196. N67 was closer to 38 introgression lines than that of IC12196. The UPGMA cluster and principal coordinate analysis (PCO) grouping showed 0.84 to 0.98 similarity values between N67 and the introgression lines. Eleven T. petropavlovskyi accessions were distinguished from introgression lines with UPGMA clusters and PCO groupings, and T. petropavlovskyi was located between the introgressions lines and IC12196. Several introgression lines resembled with T. petropavlovskyi for awning and glume length. The genetic variation among 38 introgression lines was much wider than that of T. petropavlovskyi. We concluded that T. petropavlovskyi was established by intensive selection of hybrid between T. aestivum/T. polonicum.

Variation in genome composition of blue-aleurone wheat

Key messageDifferent blue-aleurone wheats display major differences in chromosome composition, ranging from disomic chromosome additions, substitutions, single chromosome arm introgressions and chromosome translocation ofThinopyrum ponticum.AbstractAnthocyanins are of great importance for human health due to their antioxidant, anti-inflammatory, anti-microbial and anti-cancerogenic potential. In common wheat (Triticum aestivum L.) their content is low. However, elite lines with blue aleurone exhibit significantly increased levels of anthocyanins. These lines carry introgressed chromatin from wild relatives of wheat such as Thinopyrum ponticum and Triticum monococcum. The aim of our study was to characterize genomic constitutions of wheat lines with blue aleurone using genomic and fluorescence in situ hybridization. We used total genomic DNA of Th. ponticum and two repetitive DNA sequences (GAA repeat and the Afa family) as probes to identify individual chromosomes. This enabled precise localization of introgressed Th. ponticum chromatin. Our results revealed large variation in chromosome constitutions of the blue-aleurone wheats. Of 26 analyzed lines, 17 carried an introgression from Th. ponticum; the remaining nine lines presumably carry T. monococcum chromatin undetectable by the methods employed. Of the Th. ponticum introgressions, six different types were present, ranging from a ditelosomic addition (cv. Blue Norco) to a disomic substitution (cv. Blue Baart), substitution of complete (homologous) chromosome arms (line UC66049) and various translocations of distal parts of a chromosome arm(s). Different types of introgressions present support a hypothesis that the introgressions activate the blue aleurone trait present, but inactivated, in common wheat germplasm.

Development of EST-SSR markers of Ipomoea nil

Although Japanese morning glory (Ipomoea nil (L.) Roth.) has been used intensively for genetic studies, DNA markers have not been developed in Ipomoea nil sufficient to cover all chromosomes. Therefore, we conducted microsatellite (simple sequence repeats, SSR) marker development in I. nil for future genetic studies. From 92,662 expressed sequence tag (EST) sequences, 514 unique microsatellite-containing ESTs were identified. Primer pairs were designed automatically in 326 SSRs. Of 150 SSRs examined, 75 showed polymorphisms among strains. A phenogram based on the SSR genotypes revealed the genetic relation among seven Japanese morning glories from five different regions of the world and an ivyleaf morning glory (I. hederacea Jacq.). The developed SSR markers might be applicable for genetic studies of morning glories and their relatives.

Cross-species amplification of 349 melon (Cucumis melo L.) microsatellites in gherkin (Cucumis anguria L.)

Gherkin (Cucumis anguria L.), also known as West Indian gherkin, burr gherkin, and maxixe, is mainly cultivated and consumed in India, Brazil, and the United States. Marker-assisted selection (MAS) is a highly desirable tool for gherkin breeding, because gherkin cultivation generally requires time, labor, and space. However, few DNA markers for gherkin have been reported. Cross-species amplification of 349 melon (Cucumis melo L.) microsatellite primer pairs was tested on three gherkin accessions. After polymerase chain reaction optimization, 149 (42.7%) microsatellite primer pairs successfully amplified all accessions. Of the amplified primer pairs, 41 (27.5%), 64 (43.0%), and 70 (47%) showed polymorphisms between the accessions PI 147065 and PI 320052, PI 147065 and PI 364475, and PI 320052 and PI 364475, respectively. The remaining 206 primer pairs did not amplify any of the three accessions. In the polymorphic primer pairs, the correlation coefficient between repeat number and polymorphic information content values was low; therefore, it seemed unnecessary to consider it for application of repeat numbers in gherkins. Current polymorphic microsatellite primer pairs would be useful for genetic analysis, landmarks in linkage studies, studying genome structure, MAS and evolutionary ecology of Cucurbitaceae.