On the cover

The rapid identification of spontaneous mutations using a whole-genome NGS approach is important to maintain the quality of breeder seeds from the effects of deleterious mutations. We have developed such a procedure to remove deleterious mutations within a single growing season. In the upper part of the picture are the green plants and albino mutants; the middle parts include the gene interference responsible for causing albinism and the genomic differences between green and albino mutants, which is a C-insertion in the albino pool; and in the lower part are the differences in the green (left) and albino (right) leaf ultra-structures (This issue, p. 362–371).

(Y. Kishima: Research Faculty of Agriculture, Hokkaido University)

Rice stripe, a viral disease, causes widespread damage to japonica rice (Oryza sativa ssp. japonica). A rice stripe virus (RSV) bioassay revealed that many indica and japonica upland varieties exhibit resistance, whereas japonica paddy varieties are susceptible. However, the genetic background for this subspecies-dependent resistance is unclear. Herein, we focused on rice stripe resistance genes located at the Stvb locus. Three resistant alleles, Stvb-i (indica), Stvb (japonica upland), and Stvb-o (Oryza officinalis) were compared with the susceptible allele, stvb-j (japonica paddy). The expression of the resistance genes was higher than that of stvb-j. Sequence comparison revealed that the resistant and susceptible alleles had different 5ʹ-end sequences and 61-bp element(s) in the fourth intron. The insertion of an LTR-retrotransposon modified the exon 1 sequence of stvb-j. We then developed four DNA markers based on gene structure information and genotyped resistant and susceptible varieties. The LTR-retrotransposon insertion was detected only in susceptible varieties. Resistant genotypes were primarily found in indica and upland japonica, whereas paddy japonica carried the susceptible genotype. Our results characterize the genetic differences associated with RSV resistance and susceptibility in O. sativa and provide insights on the application of DNA markers in rice stripe disease management.

The timing of heading is largely affected by environmental conditions. In wheat, Vrn-1 and Ppd-1 have been identified as the major genes involved in vernalization requirement and photoperiod sensitivity, respectively. To compare the effects of Vrn-1 and Ppd-1 alleles on heading time under different environments, we genotyped Vrn-1 and Ppd-1 homoeologues and measured the heading time at Morioka, Tsukuba and Chikugo in Japan for two growing seasons. A total of 128 Japanese and six foreign varieties, classified into four populations based on the 519 genome-wide SNPs, were used for analysis. Varieties with the spring alleles (Vrn-D1a or Vrn-D1b) at the Vrn-D1 locus and insensitive allele (Hapl-I) at the Ppd-D1 locus were found in earlier heading varieties. The effects of Vrn-D1 and Ppd-D1 on heading time were stronger than those of the other Vrn-1 and Ppd-1 homoeologues. Analysis of variance revealed that heading time was significantly affected by the genotype-environment interactions. Some Vrn-1 and Ppd-1 alleles conferred earlier or later heading in specific environments, indicating that the effect of both alleles on the timing of heading depends on the environment. Information on Vrn-1 and Ppd-1 alleles, together with heading time in various environments, provide useful information for wheat breeding.

Improvement of preharvest sprouting (PHS) resistance is an important objective in the breeding of durum wheat (Triticum turgidum ssp. durum (Desf.) Husn.) in Japan, where the harvest timing overlaps with the rainy season. In a previous study, we showed that an R-gene associated with red seed color was the most effective at promoting PHS resistance in durum wheat. However, red-seeded durum wheat is not popular because it discolors pasta. Here, to improve PHS resistance without the R-gene, we introduced a PHS resistance allele of MOTHER OF FT AND TFL 1 (MFT) and a mutated ABA 8ʹ-hydroxylase (ABA8ʹOH1-A), which is involved in abscisic acid (ABA) catabolism, singly or together into white-seeded durum wheat. The introduction of both genes reliably and stably improved PHS resistance under all tested conditions. Modification of ABA catabolism might be an effective way to improve PHS resistance in durum wheat. Our findings will contribute to improved PHS resistance in breeding for white-seeded durum wheat.

Spontaneous mutations are stochastic phenomena that occur in every population. However, deleterious mutated allele present in seeds distributed to farmers must be detected and removed. Here, we eliminated undesirable mutations from the parent population in one generation through a strategy based on next-generation sequencing (NGS). This study dealt with a spontaneous albino mutant in the ‘Hinohikari’ rice variety grown at the Miyazaki Comprehensive Agricultural Experiment Station, Japan. The incidence of albinism in the population was 1.36%. NGS analysis revealed the genomic basis for differences between green and albino phenotypes. Every albino plant had a C insertion in the Snow-White Leaf1 (SWL1) gene on chromosome 4 causing a frameshift mutation. Selfing plants heterozygous for the mutant allele, swl1-R332P, resulted in a 3:1 green/albino ratio, confirming that a single recessive gene controls albinism. Ultrastructural leaf features in the swl1-R332P mutants displayed deformed chlorophyll-associated organelles in albino plants that were similar to those of previously described swl1 mutants. Detection of the causative gene and its confirmation using heterozygous progenies were completed within a year. The NGS technique outlined here facilitates rapid identification of spontaneous mutations that can occur in breeder seeds.

Japanese soil-borne wheat mosaic virus (Furovirus) is a damaging pathogen of wheat and barley. This virus can survive in the soil for several decades, so the deployment of resistant cultivars represents the only practical control measure. Here, a genetic analysis has identified two regions of the barley genome—one on chromosome 2H and the other on chromosome 3H—as harboring gene(s) encoding resistance to this virus. The joint presence of both loci, termed Jmv1 and Jmv2, made the plants essentially immune, with resistance being dominant over susceptibility at each locus. Phylogenetic analysis showed that the virus is not closely related to the type Furovirus species Soil-borne wheat mosaic virus. There was a difference between the RNA1- and RNA2-based phylogenies of the virus species in Furovirus implying the independent segregation of the virus subgenomes.

Rice flour is useful as a substitute for wheat flour, however, to obtain fine flour, millers need special milling facilities, which increase the cost of milling. To reduce the milling cost, we developed a floury mutant line by irradiating gamma-rays to dry seeds of the japonica cultivar ‘Hoshinoyume’. The line was registered as a new cultivar, ‘Hoshinoko’. Genetical analysis of the floury trait was conducted using an F₂ population derived from a cross between ‘Hoshinoko’ and ‘Corbetti’ (a japonica rice cultivar with normal endosperm), which indicated the involvement of a single recessive gene located near the RM163 marker on the long arm of rice chromosome 5, flanking flo4 identified by Kang et al. (2005). Sequence analysis of flo4 showed a two-bp (CA) insertion in the eighth exon of in ‘Hoshinoko’ compared to that of ‘Hoshinoyume’, which led to a frameshift mutation. The CAPS-based genotype of flo4 gene completely correlated to the phenotype of endosperm in two populations. This CAPS marker could be helpful for rice breeders to develop new cultivars harboring floury endosperm of the flo4-303 gene.