On the cover

Purple corn with a high anthocyanin content is a promising genetic resource due to the possibility of mitigating oxidative stress, which reduces milk production in dairy cows. The Peruvian cultivar ‘JC072A’ (left) is one of purple corn and shows many organs with anthocyanin pigmentation such as leaf sheath (upper center), bract leaf (upper center), plant ear (upper center and right), grain and cob (lower center). The genetic information about anthocyanin pigmentation in JC072A assists maize breeders in developing the molecular markers for the effective breeding of F₁ hybrids of purple corn (This issue, p. 582–586).

(J. Yonemaru: NARO, Institute of Crop Science)

The melon fly, Bactrocera cucurbitae (Tephritidae: Diptera) is an important pest of snapmelon (Cucumis melo var. momordica), leading to significant losses in yield in the hot arid agro-climate of India. The accessions IC-430190 (11.21%), DKS-AHS 2011/4 (14.97%) and DKS-AHS 2011/3 (18.57%) were found to be novel resistant accessions against melon fly, B. cucurbitae infestation. Free amino acid and total soluble solid (TSS) were in positive correlation with percent fruit infestation whereas phenols, tannin, total alkaloids and flavonoid contents had significant negative correlation with percent fruit infestation. The percent fruit infestation had significant positive correlation with fruit length, fruit diameter and flesh thickness and negative correlation with length of ovary pubescence, rind hardness at immature stage, rind hardness at mature stage and pericarp thickness. Based on Kaiser Normalization method, two principal components (PCs) were extracted explaining cumulative variation of 82.80% in melon fly infestation. PC1 explained 53.41% of the variation while PC2 explained 29.39% of variation. The flavonoid, total alkaloid, tannins, phenols content, length of ovary pubescence and rind hardness were the novel antibiosis and antixenotic characters found in snapmelon resistant melon fly, B. cucurbitae and therefore, could be used as marker traits in plant breeding programs to select resistant accessions.

Lodging in crops seriously restricts plant growth and grain production. The genetic modification of cell walls to enhance plant mechanical strength has been suggested as a promising approach toward improving lodging resistance. However, because of the complexity of the plant cell wall, the exact effects of its polymers on plant lodging resistance remain elusive. To address this issue, we performed large-scale analyses of a total of 56 rice (Oryza sativa L.) varieties that displayed distinct cell wall component and lodging index. Lignin was identified as the key cell wall polymer that positively determines lodging resistance in rice. Correlation analysis between cell wall composition and plant morphological characteristics revealed that lignin enhanced rice lodging resistance by largely increasing the mechanical strength of the basal stem and reducing plant height. Further characterization of four representative rice varieties, ShenNong9903, YanJian218, KongYu131, and ShenNongK33, displaying varied levels of lodging resistance, revealed the multiple candidate genes (PAL, CoMT, 4CL3, CAD2, CAD7 and CCR20) responsible for increasing lignin level. Hence, our results demonstrate that the high lignin level in the cell wall predominately improves lodging resistance and suggest target genes for the genetic modification of lignin towards breeding rice with high lodging resistance.

Oryza meridionalis is a potential source for improving Asian cultivated rice O. sativa via direct hybridization and backcrossing. However, hybrid sterility between O. sativa and O. meridionalis is the main barrier of reproduction hindering the transfer of favorable genes from O. meridionalis to O. sativa. To investigate the nature of hybrid sterility between O. sativa and O. meridionalis, three accessions of O. meridionalis were used as male parents to cross Dianjingyou 1, an O. sativa subsp. japonica cultivar following the backcross with the recurrent parent of Dianjingyou 1. Twenty pollen sterility NILs (BC₆F₁) were obtained and genotyped by using simple sequence repeat (SSR) markers distributed across the 12 rice chromosomes. The heterozygous markers were employed to genotype the corresponding segregation populations for mapping sterility genes. As a result, five novel loci for pollen sterility between O. sativa and O. meridionalis were identified and designated as S51(t), S52(t), S53(t), S54(t) and S55(t), respectively. The genetic behavior of five novel loci followed one-locus allelic interaction model. The disharmonious interaction between Asian cultivated rice allele and wild relative allele led to the partial or full abortion of male gametes for one parent allele in the heterozygotes. These results will be useful for elucidating the mechanism of interspecific hybrid sterility and further utilizing favorable genes from O. meridionalis for enhancement of rice breeding.

Low molecular weight glutenin subunits (LMW-GS) play an important role in determining the bread-making characteristics of dough in the end-use quality of wheat. In this study, A total of 149 worldwide-originated durum wheat were used to analyze the composition of LMW-GS using MALDI-TOF-MS. Based on the allelic variation of glutenin subunits, the genetic diversity was evaluated for the 149 durum wheat. Five types of alleles were identified at the Glu-A3 locus with Glu-A3e, Glu-A3a/c, Glu-A3f, Glu-A3d and Glu-A3b accounting for 43.0%, 16.1%, 12.8%, 10.1% and 7.4 % of the accessions, respectively. Five types of alleles were identified at the Glu-B3 locus: Glu-B3d (60.4%), Glu-B3b (6.0%), Glu-B3c (6.0%), Glu-B3h (2.7%) and Glu-B3f (0.7%). Two novel alleles encoding abnormal subunits 40500 Da and 41260 Da were identified at the Glu-A3 and Glu-B3 loci, respectively. Further studies are needed to match these novel alleles to previously discovered novel alleles. Moreover, the genetic diversity analysis indicated that great genetic variation existed in durum wheat among encoding loci of glutenin subunits, released periods of varieties and different geographical origins. The results provide more important information of potential germplasm for the improvement of durum wheat and common wheat.

Rhododendron possesses valuable horticultural and medicinal properties. However, the genetic studies have been hindered due to the lack of genetic markers. Based on RNA-seq, large-scale molecular markers were developed from four Rhododendron species endemic to Dabie Mountains (central China): R. fortunei (5.25 Gb; SSRs, 12,756, one/2.37 kb, 147 types; SNPs, 38,313; InDels, 3,174), R. simsii (5.80 Gb; SSRs, 13,294, one/2.58 kb, 167 types; SNPs, 136,590; InDels, 6,258), R. mariesii (6.53 Gb; SSRs, 15,724, one/2.51 kb, 170 types; SNPs, 44,942; InDels, 4,126), and R. molle (4.35 Gb; SSRs, 10,214, one/2.49 kb, 110 types; SNPs, 77,829; InDels, 3,416). Di-nucleotide repeats were the main type (59.126%–64.314%), and AG/CT repeat (55.18%–61.22%) was the most. In particular, 89 species-specific types had been found. Furthermore, C:G→T:A mutation was the main SNP type (30.475%–34.99%). However, C:G→G:C mutation was the least type in R. fortunei, while T:A→G:C mutation was the least in the other three species. InDels with length of 3 nt was most in R. fortunei, but 1 nt InDels were the main type in the other three species. Twelve microsatellite markers developed from R. simsii reveled high genetic diversity in the four populations, and heterozygote excess was observed. This research would benefit the genetic study, molecular marker-assisted selection, and breeding studies in Rhododendron species.

The international cacao collection in CATIE, Costa Rica contains nearly 1200 accessions of cacao, mainly from the center of genetic diversity of this species. Among these accessions, the United Fruit clones (UF clones) were developed by the United Fruit Company in Costa Rica, and they represent one of the earliest groups of improved cacao germplasm in the world. Some of these UF clones have been used as key progenitors for breeding resistance/tolerance to Frosty Pod and Black Pod diseases in the Americas. Accurate information on the identity and background of these clones is important for their effective use in breeding. Using Single Nucleotide Polymorphism (SNP) markers, we genotyped 273 cacao germplasm accessions including 44 UF clones and 229 reference accessions. We verified the true-to-type identity of UF clones in the CATIE cacao collection and analyzed their population memberships using maximum-likelihood-based approaches. Three duplicate groups, representing approximately 30% of the UF clones, were identified. Both distance- and model-based clustering methods showed that the UF clones were mainly composed of Trinitario, ancient Nacional and hybrids between ancient Nacional and Amelonado. This result filled the information gap about the UF clones thus will improve their utilization for cacao breeding.

Manipulating the genetic control of plant height is essential in soybean breeding to increase yield through the enlargement of the plant size while preventing lodging. A Japanese soybean germplasm, Y2, has distinctively shorter inter-node lengths than those of recently developed Japanese cultivars and is expected to provide new variation to prevent lodging. A quantitative trait loci (QTL) analysis for plant height-related traits was conducted using F₂ individuals derived from a cross between the elite Japanese cultivar Fukuyutaka and Y2. A major QTL for average inter-node length (AIL) and plant height was identified on chromosome 13 and named qSI13-1 (QTL for short inter-node on chromosome 13). The Y2 allele of qSI13-1 was partially dominant for plant height. qSI13-1 exhibited no effect on either days to flowering or number of main stem nodes. The AILs and plant heights of the near-isogenic lines containing the Y2 allele of qSI13-1 in the genetic background of Fukuyutaka were significantly less than those of Fukuyutaka. No significant differences between the near-isogenic lines and Fukuyutaka were observed for seed yield and flowering date, indicating that qSI13-1 will be useful in developing cultivars with short plant heights without having negative effects on yield potential and days to flowering.

Enhancing salt stress tolerance is a key strategy for increasing global food production. We previously found that long-term salinity stress significantly reduced grain fertility in the salt-sensitive barley (Hordeum vulgare) accession, ‘OUC613’, but not in the salt-tolerant accession, ‘OUE812’, resulting in large differences in grain yield. Here, we examined the underlying causes of the difference in grain fertility between these accessions under long-term treatment with 150 or 200 mM NaCl from the seedling stage to harvest and identified quantitative trait loci (QTLs) for maintaining grain fertility. In an artificial pollination experiment of the two accessions, grain fertility was significantly reduced only in OUC613 plants produced using pollen from plants grown under NaCl stress, suggesting that the low grain fertility of OUC613 was mainly due to reduced pollen fertility. Using QTL-seq combined with exome-capture sequencing and composite interval mapping of recombinant inbred lines derived from a cross between OUE812 and OUC613, we identified a QTL (qRP-2Hb) for grain fertility on chromosome 2H. The QTL region includes two genes encoding an F-box protein and a TIFY protein that are associated with male sterility, highlighting the importance of this region for maintaining grain fertility under salt stress.

Variegation is a frequently observed genetic phenomenon in landscaping. In this study, an ethyl methanesulfonate induced variegated leaf (Csvl) mutant in cucumber (Cucumis sativus L.) was identified. The Csvl mutant displayed green-yellow-white variegation phenotype throughout the whole growth cycle, while the leaf of wild type plants was normal green. The photosynthetic pigment contents and photosynthetic parameters of Csvl was significantly lower than wild type. The cytology observation results showed that the mesophyll cells of Csvl mutant contained defective chloroplasts. Genetic analysis indicated that variegated leaf phenotype was monogenic recessive inheritance. MutMap and genotyping results revealed that Csa6G405290 (Cscs), encoding chorismate synthase, was the candidate gene for variegated leaf mutant in cucumber. The expression level of Cscs was similar between wild type and variegated leaf mutant leaves. Transcriptome profile analysis of leaves of Csvl mutant identified 183 candidate genes involved in variegated leaf development in cucumber, including genes that encode heat shock protein, zinc finger protein. Cscs may regulate variegated leaf in cucumber by interacting with these genes. In a word, these results revealed that Cscs might regulate the variegated leaf phenotype in cucumber.

Purple corn is a maize variety (Zea mays L.) with high anthocyanin content. When purple corn is used as forage, its anthocyanins may mitigate oxidative stresses causing lower milk production in dairy cows. In this study, we analyzed quantitative trait loci (QTLs) for anthocyanin pigmentation of maize organs in an F₂ population derived from a cross between the Peruvian cultivar ‘JC072A’ (purple) and the inbred line ‘Ki68’ (yellowish) belonged to Japanese flint. We detected 17 significant QTLs on chromosomes 1–3, 6, and 10. Because the cob accounts for most of the fresh weight of the plant ear, we focused on a significant QTL for purple cob on chromosome 6. This QTL also conferred pigmentation of anther, spikelet, leaf sheath, culm, and bract leaf, and was confirmed by using two F₃ populations. The gene Pl1 (purple plant 1) is the most likely candidate gene in this QTL region because the amino acid sequence encoded by Pl1-JC072A is similar to that of an Andean allele, Pl-bol3, which is responsible for anthocyanin production. The markers designed for the Pl1 alleles will be useful for the breeding of F₁ lines with anthocyanin pigmentation in cobs.

Spelt wheat (Triticum aestivum subsp. spelta), a subspecies of common wheat, is a genetic resource for the breeding of bread wheat (T. aestivum subsp. aestivum); however, genetic analyses of agronomic traits in bread wheat × spelt crosses are insufficient. Here, we conducted QTL analysis in the recombinant inbred lines from a bread wheat × spelt cross. In addition to the major Q locus, QSpd.obu-4D was detected with the spelt allele conferring a higher spikelet density than the bread wheat allele. The effect of QSpd.obu-4D was evident in the presence of the Q allele of bread wheat, suggesting that this variation might be cryptic in spelt wheat with the q allele. Two QTLs with stable effects were identified for grain length, one of which (QGl.obu-1A) has never been detected in a bread wheat × spelt cross. The spelt wheat allele at QHt.obu-7B conferring later heading was identified in the Vrn-B3 region and could be a novel gene source for modifying heading time. Furthermore, QGi.obu-2B, responsible for low grain dormancy of spelt wheat, was detected. Further exploration and identification of useful QTLs could accelerate the utilization of spelt wheat as a genetic resource for bread wheat breeding programs.

High-density genetic linkage maps are particularly important for quantitative trait loci (QTL) mapping, genome assembly, and marker-assisted selection (MAS) in plants. In this study, a high-density genetic linkage map of sunflower (Helianthus annuus L.) was constructed using an F₂ population generated from a cross between Helianthus annuus L. ‘86-1’ and ‘L-1-OL-1’ via specific-locus amplified fragment sequencing (SLAF-seq). After sequence preprocessing, 530.50 M reads (105.60 Gb) were obtained that contained a total of 343,197 SLAFs, of which 39,589 were polymorphic. Of the polymorphic SLAFs, 6,136 were organized into a linkage map consisting of 17 linkage groups (LGs) spanning 2,221.86 cM, with an average genetic distance of 0.36 cM between SLAFs. Based on this high-density genetic map, QTL analysis was performed that focused on four sunflower phenotypic traits: oleic acid content (OAC), plant height (PH), head diameter (HD), and stem diameter (SD). Subsequently, for these four traits eight QTLs were detected that will likely be useful for increasing our understanding of genetic factors underlying these traits and for use in marker-assisted selection (MAS) for future sunflower breeding.

Grain number per panicle is a major component of rice yield that is typically controlled by many quantitative trait loci (QTLs). The identification of genes controlling grain number per panicle in rice would be valuable for the breeding of high-yielding rice. The Oryza glaberrima chromosome segment substitution line 9IL188 had significantly smaller panicles compared with the recurrent parent 9311. QTL analysis in an F₂ population derived from a cross between 9IL188 and 9311 revealed that qgnp7(t), a major QTL located on the short arm of chromosome 7, was responsible for this phenotypic variation. Fine mapping was conducted using a large F₃ population containing 2250 individuals that were derived from the F₂ heterozygous plants. Additionally, plant height, panicle length, and grain number per panicle of the key F₄ recombinant families were examined. Through two-step substitution mapping, qgnp7(t) was finally localized to a 41 kb interval in which eight annotated genes were identified according to available sequence annotation databases. Phenotypic evaluation of near isogenic lines (NIL-qgnp7 and NIL-qGNP7) indicated that qgnp7(t) has pleiotropic effects on rice plant architecture and panicle structure. In addition, yield estimation of NILs indicated that qGNP7(t) derived from 9311 is the favorable allele. Our results provide a foundation for isolating qgnp7(t). Markers flanking this QTL will be a useful tool for the marker-assisted selection of favorable alleles in O. glaberrima improvement programs.

Wild rice, Oryza rufipogon, is a genetic resource that can be used to improve cultivated rice, but its populations are now decreasing in terms of both size and number. Extensive research on wild rice has been conducted in Thailand, where two in situ conservation sites have been preserved in natural areas where perennial wild rice predominates. The genetic structure of wild rice populations was investigated by examining both the chloroplast and nucleus genomes at sites of in situ conservation site in Thailand. One accession from an in situ-conserved site was re-sequenced against the chloroplast genome of O. sativa cv. ‘Nipponbare’ to develop chloroplast insertion/deletion (cpINDEL) markers. These cpINDEL markers revealed unique maternal lineages in the in situ-conserved populations upon comparison with other Asian wild rice accessions. Diverse genetic variation was also detected with SSR markers throughout the genome. Three populations differed from each other and also within single populations. The sub-populations within an in situ-conserved population showed a complex population structure due to their multiple maternal lineages and relatively higher number of haplotypes when they maintained a relatively large population size. Such a heterogeneous population would serve as a unique gene pool for rice breeding.

Kernel moisture content at harvest stage (KMC) is an important factor affecting maize production, especially for mechanical harvesting. We investigated the genetic basis of KMC using an association panel comprising of 144 maize inbred lines that were phenotypically evaluated at two field trial locations. Significant positive or negative correlations were identified between KMC and a series of other agronomic traits, indicating that KMC is associated with other such traits. Combining phenotypic values and the Maize SNP3K Beadchip to perform a genome-wide association study revealed eight single nucleotide polymorphisms (SNPs) associated with KMC at P ≤ 0.001 using a mixed linear model (PCA+K). These significant SNPs could be converted into five quantitative trait loci (QTLs) distributed on chromosomes 1, 5, 8, and 9. Of these QTLs, three were co-localized with genomic regions previously reported. Based on the phenotypic values of the alleles corresponding to significant SNPs, the favorable alleles were mined. Eight maize inbred lines with low KMC and harboring favorable alleles were identified. These QTLs and elite maize inbred lines with low KMC will be useful in maize breeding.

To induce potato variants with enhanced resistance to common scab disease that retain the desirable agronomic traits of the original cultivars, we used a cell culture technique that employs thaxtomin A, the primary phytotoxin that induces scab symptoms. We induced 24 variants from the potato cultivar ‘Saya-akane’, developed in Japan, and selected two with enhanced resistance to the disease by growing them in planters with bacteria-inoculated soil and in a field infested with the disease. We also examined toxin tolerance in micro-tubers of variants that showed a lower degree or percentage of infection in the glasshouse screening, and found no significant difference relative to the original cultivar. To clarify the effect of using thaxtomin A, we examined the efficiency of induction of the potential enhanced resistance by comparing the degree of infection among variants grown in planters with inoculated soil. We observed no significant difference between variants induced on culture medium with and without the toxin. These results suggest that the effect of using the toxin as a positive selection agent is restrictive and that most resistance-enhancing mutations are induced by the cell culture procedure itself.