In soybean, chilling temperatures negatively affect seed appearance by causing seed coat discoloration around the hilum region, which is called cold-induced discoloration (CD). The Ic marker, the DNA marker associated with CD tolerance, has been developed under phytotron conditions. In this issue, we assessed the effect of the Ic marker on CD tolerance under a cool field environment (main photo). The average pigment index of the cultivar ‘Toyomizuki’ (I allele, upper first left panel) was significantly higher than that of the near-isogenic line of ‘Toyomizuki’ (Ic allele, upper second left panel) in the field test (This issue, p. 521–528).
(N. Yamaguchi: Hokkaido Research Organization Tokachi Agricultural Experiment Station)
Cacao (Theobroma cacao L.) is considered a key crop in Colombian social programs aiming at alleviating rural poverty, promoting peace in post-conflict regions and, replacing crops used for illicit purposes. Colombia is thought to be part of the center of origin of cacao; several germplasm collecting expeditions have been implemented, dating back to the 1940s. Despite that history, the first breeding program based on creating, selecting, and releasing full-sib progenies made extensive use of accessions introduced from other countries as parents. A new breeding strategy was adopted in the 1990s, based on mass selection of promising trees (high-yield and disease-resistant) in farmers’ fields, resulting in the selection of clones released to farmers as planting material. In 2012, a new strategy, Recurrent Selection, was adopted by the Colombian Corporation for Agricultural Research, Agrosavia, based on the development of improved populations and allowing the selection of clones at the end of each cycle of recombination. The use of molecular markers is being integrated into this program in order to assist breeders in selecting material. This review provides details about the history and perspectives of the cacao breeding program in Colombia.
Rice breeding programs in Hokkaido over the past 100 years have dramatically increased productivity and improved the eating quality of rice. Commercial varieties with high yield and good eating quality, such as Kirara 397, Hoshinoyume, and Nanatsuboshi, have been continuously registered since 1990. Furthermore, varieties with better eating quality using Wx1-1, which reduces amylose content to improve the taste of sticky rice, such as Oborozuki and Yumepirika, were registered in 2006 and 2008, respectively. However, to the best of our knowledge the genomic changes associated with these improvements have not been determined. Better understanding of the relationships between DNA sequences and agricultural traits could facilitate rice breeding programs in Hokkaido. Marker-assisted selection (MAS), which can select the plants with chromosomal regions tagged with DNA markers for desirable traits, is an advanced technology to manage genetic improvements. Here, we summarize the current states of MAS in rice breeding programs in Hokkaido before huge data sets of genome sequences using next-generation sequencing technology come into practical use in rice breeding programs.
A clonal line of Camellia taliensis, ‘Taliensis-akeme’ has a recessive caffeine-less gene. To accelerate breeding of caffeine-less tea cultivars using this gene, DNA markers are indispensable for selecting heterozygotes that do not show a caffeine-less phenotype as parental lines. Therefore, we tried to determine the sequence of the six tea caffeine synthase (TCS) genes to search for polymorphisms and to prepare one of the TCS genes as a selection marker. Six TCS genes and the caffeine-less trait were mapped on the reference linkage map of tea. Strong linkage between the caffeine-less phenotype and TCS1 indicate that it is a promising candidate as a causative gene of the caffeine-less trait. We decided to use a three-nucleotide insertion in TCS1 that can be distinguished by sequencing as a selection marker named ‘CafLess-TCS1’. Caffeine-less individuals appeared in the progeny population of caffeine-less heterozygous individuals selected using ‘CafLess-TCS1’. These results confirmed that the developed ‘CafLess-TCS1’ will be an effective selection marker for breeding of caffeine-less tea cultivars.
Sake-brewing cultivars among varieties of Japanese rice (Oryza sativa L.) have traits adapted to the sake-brewing process, such as a high white-core expression rate (WCE). Our previous study detected putative quantitative trait loci (QTLs) associated with a high WCE derived from Yamadanishiki, a popular brewing rice cultivar. Because the occurrence of white-core grains depends on air temperature and the position of the grain on the panicle, phenotyping of WCE must consider these variable conditions. In this study, qWCE6, a QTL for the WCE on chromosome 6, was validated for the first time, and the phenotyping method examined for its suitability in fine-mapping. A clear tendency towards high WCE was observed in late-heading substituted lines which headed under low daily mean temperature at the experimental location. White-core grains were often expressed by the primary spikelets on the upper panicle, producing a high percentage of superior grains. The segregating population for qWCE6 in late heading revealed a distinct difference in WCE between the Koshihikari and Yamadanishiki homozygous alleles at qWCE6 as determined from that locality. Further, two insertion/deletion markers were developed for the marker-assisted selection of qWCE6. Our results will be useful for informing the breeding of sake-brewing rice cultivars.
Apple is an economically important crop, and various approaches to genetic analysis in breeding programs have been attempted, including the production of doubled haploid (DH) lines, which are genetically homozygous. In this study, we used a DH line for QTL analyses, for the first time in a fruit tree, expecting it to simplify the analysis of the inheritance of quantitative traits and thus to enhance QTL detection power. Using an F1 population from ‘Prima’ × ‘Apple Chukanbohon 95P6’ (DH), we constructed a genetic map of ‘Prima’, and identified 19 QTLs for 13 traits. These QTLs had comparatively high LOD scores and explained a large part of the variation of the phenotypes. In particular, acidity, juice browning, and skin splitting clearly segregated at a 1:1 ratio, consistent with the segregation of the alleles at the detected QTLs in linkage group 16; these traits appeared to be regulated by single genes, despite general consideration that they are quantitative traits. Using this simple genetic composition of the F1 population, we concluded that the skin splitting of apple fruit has recessive inheritance, and that the allele for splitting is tightly linked with those for high acidity and low juice browning in ‘Prima’.
Yield improvement is a top priority for maize breeding. Kernel size and weight are important determinants of maize grain yield. In this study, a recombinant inbred line (RIL) population and an association panel were used to identify quantitative trait loci (QTLs) for four maize kernel-related traits: kernel length, width, thickness and 100-kernel weight. Twenty-seven QTLs were identified for kernel-related traits across three environments and the best linear unbiased predictions (BLUPs) of each trait by linkage analysis, and four QTLs were stably detected in more than two environments. Additionally, 29 single nucleotide polymorphisms (SNPs) were identified as significantly associated with the four kernel-related traits and BLUPs by genome-wide association study, and two loci could be stably detected in both environments. In total, four QTLs/SNPs were co-associated with various traits in both populations. Using combined-linkage analysis and association mapping, PZE-101066560 on chromosome 1, associated with kernel width and with 100-kernel weight in the association panel, was co-localized within the QTL interval of qKW1-3 for kernel width in the RILs. Two annotated genes in the candidate region were considered as potential candidate genes. The QTLs and candidate genes identified here will facilitate molecular breeding for grain yield improvement in maize.
As glutamate dehydrogenases (GDHs) of microorganisms usually have higher affinity for NH4+ than do those of higher plants, it is expected that ectopic expression of these GDHs can improve nitrogen assimilation in higher plants. Here, a novel NADP(H)-GDH gene (TrGDH) was isolated from the fungus Trichurus and introduced into rice (Oryza sativa L.). Investigation of kinetic properties in vitro showed that, compared with the rice GDH (OsGDH4), TrGDH exhibited higher affinity for NH4+ (Km = 1.48 ± 0.11 mM). Measurements of the NH4+ assimilation rate demonstrated that the NADP(H)-GDH activities of TrGDH transgenic lines were significantly higher than those of the controls. Hydroponic experiments revealed that the fresh weight, dry weight and nitrogen content significantly increased in the TrGDH transgenic lines. Field trials further demonstrated that the number of effective panicles, 1,000-grain weight and grain weight per plant of the transgenic lines were significantly higher than those of the controls, especially under low-nitrogen levels. Moreover, glutelin and prolamine were found to be markedly increased in seeds from the transgenic rice plants. These results sufficiently confirm that overexpression of TrGDH in rice can improve the growth status and grain weight per plant by enhancing nitrogen assimilation. Thus, TrGDH is a promising candidate gene for maintaining yields in crop plants via genetic engineering.
The green rice leafhopper (GRH), Nephotettix cincticeps Uhler, is a major insect pest of cultivated rice, Oryza sativa L., throughout the temperate regions of East Asia. GRH resistance had been reported in the wild species Oryza nivara but genetic basis of GRH resistance in wild rice accession has not been clarified. Here, we found a major QTL, qGRH4.2, on chromosome 4 conferred GRH resistance with 14.1 of the logarithm of odds (LOD) score explaining 67.6% of phenotypic variance in the BC1F1 population derived from a cross between the susceptible japonica cultivar ‘Taichung 65’ (T65) and O. nivara accession IRGC105715. qGRH4.2 has been identified as GRH6 between the markers RM5414 and C60248 in a BC3F2 population derived from two BC3F1 plants resistant to GRH. In a high-resolution mapping, the GRH6 region was delimited between the markers G6-c60k and 7L16f, and corresponded to an 31.2-kbp region of the ‘Nipponbare’ genome. Understanding the genetic basis of GRH resistance will facilitate the use of GRH resistance genes in marker-assisted breeding in rice.
Fruit weight (FW) and shelf life (SL) are important traits in commercial fresh market tomatoes. A tomato RIL population was developed by antagonistic and divergent selection for both traits from an interspecific cross between the Solanum lycopersicum L. cv. “Caimanta” and the S. pimpinellifolium L. accession “LA0722”. The objective of this work was to evaluate phenotypic and genetic components for FW and SL. Phenotypic data from RILs were collected during 3-year trials. Sixteen SSR, 62 InDels developed based on the genome sequences of “Caimanta” and “LA0722”, and four functional markers for fruit size genes were used. FW and SL had a significant genetic variability, and both traits showed a genotype by year interaction. Genome-wide molecular characterization of the population demonstrated that is genetically structured according to FW. Marker data was used to study changes on allelic frequencies at loci between the phenotypic extreme group of RILs for FW and SL. Twenty four markers were associated to FW, the LC gene in chromosome 2 and other six markers in chromosomes 1, 2, 6, and 11 presented the most significant associations. Finally, we reported three new genomic regions located on chromosomes 9, 10 and 12 that underlie SL in tomato.
Although cultivation of hybrid rice varieties has been increasing, there are risks that high levels of cadmium (Cd) will accumulate in grain when such rice is grown in Cd-polluted environments. To produce Cd-safe hybrid rice, one practical approach is the generation of low Cd-accumulating parental lines. In two-line hybrid breeding, thermosensitive genic male sterile (TGMS) lines function as female parents to yield hybrid seeds. Recently, Cd accumulation-related genes have been identified; however, the effect of these genes on Cd accumulation in the grains of TGMS lines has yet to be reported. Here, 174 TGMS lines were selected for Cd accumulation phenotyping, and 30 TGMS lines, including 15 stable low-Cd and 15 high-Cd lines, were selected for single-nucleotide polymorphism (SNP) genotyping and association analysis. Association studies were conducted to identify the relationship between Cd accumulation and variable sites within seven candidate Cd-associated genes using logistic models. Nine sequence variant sites in four of the candidate genes were found to be significantly associated with Cd accumulation, two of which in OsNRAMP1 and OsNRAMP5 are low-Cd favorable variants, explaining 46.4% and 22.6% of the phenotypic variation, respectively. These loci could be developed as new molecular markers for identification of Cd accumulation characteristics and low-Cd marker-assisted breeding.
In order to solve the difficult problem of the outer shell covering in the processing of Tartary buckwheat, we conducted a genetic analysis in segregating F2 and F3 populations derived from a hybrid between ‘Yunqiao No. 1’ and ‘Rice buckwheat’, and the F3 population was used to analyze the phenotypic and genetic correlation among the traits. The results showed that the variety with a value of trait for seed shell thickness over 0.20 mm is a hard-shelled type (The thick shell type = seed shell rate > 20%), and that with a value of trait for seed shell thickness below 0.15 mm is a easily-shelled type (The thin shell type = seed shell rate < 20%), while that with a value of trait for seed shell thickness ranging from 0.15 mm to 0.20 mm is a hard-shelled type or easily-shelled type. In addition, alleles for traits of number of seed per plant and total seed weight per plant have larger dominance variance relative to their additive variance, indicating that genes controlling these traits have larger dominant effects, it is not suitable for the selection of single plant in offspring plants at the early stage of development, because these traits do not show up then. The alleles for traits of 1000 kernel weight and seed shell thickness have larger additive variance relative to their dominant variance, indicating that genes governing these traits have greater additive effects, with which the single plant can be selected in the progeny at the early stage of development. Although, the value of seed shell thickness has been shown to correlated positively with that of 1000 kernel weight, almost all the seeds of easily-shelled type are those with thin shell. However, ideal single plants with easily-shelled trait are those with intermediate phenotypes of seed shell thickness and 1000 kernel weight, by which the traits of large number of seeds per plant and total seed weight per plant could be selected. In the progeny population of this study, there were excellent single plants with high-yield and easily-shelled traits, of which the value of seed shell thickness was 0.17 mm (0.15 mm to 0.20 mm), the value of 1000 kernel weight was 14 g, the value of number of seeds per plant was 1137 and value of total seed weight per plant was 15.9 g. The results showed that taking the hybrid combinations of easily-shelled trait with the trait of seed shell thickness was the most effective selection indexes to breed the high-yield buckwheat varieties with the trait of easy shelling.
To evaluate genetic diversity and genetic structure of wild rice (Oryza rufipogon) populations in Myanmar, seven research sites were selected based on various ecological conditions. A large number of samples under natural growth conditions were collected and studied using six simple sequence repeats (SSRs) and two chloroplast DNA markers. A total of 77 alleles were detected from 1559 samples over six SSR loci. The mean number of alleles per population ranged from 3.167 to 8.667, and the mean expected heterozygosity ranged from 0.140 to 0.701. Wild rice populations survived under various environmental conditions and retained different levels of genetic diversity. The large number of samples was effective to confirm the spatial genetic structure of wild rice populations in a relatively small area. Regarding chloroplast DNA polymorphisms, four populations possessed only one pattern, while the other three showed two or five combinations of haplotypes, even within the same population. Additionally, the existence of a new genotype was revealed. Considerable variations in chloroplast DNA exist in the wild rice populations of Myanmar. A high proportion of genetic variation was detected within, rather than among, populations. To ensure maintenance of allelic diversity, it is advisable to preserve many individuals from a large population.
The physical properties of various white bread doughs made from the flours of ‘Harunoakebono’ and 10 genotypes of its near-isogenic lines with different compositions of high molecular weight glutenin subunit (HMWGs) were measured with the Creep method based on a Maxwell–2–element model. The expansion stress in the proofing process of various doughs was obtained by a numerical calculation method. The results indicated that doughs with high elastic characteristics, namely large relaxation time (τ0) and regularity coefficient of viscosity (ηN), have high dough stress throughout the proofing process and high stress at the proofing end (σend) and conversely, the low elastic dough with the small τ0 and ηN has the completely opposite tendency. This study also showed that there are significantly high correlations between the calculated σend and bread-making quality (BMQ) such as gas retention of dough and specific loaf volume (SLV). These results showed that BMQ, represented by SLV, of various white bread doughs were greatly influenced by the dough’s physical properties, especially τ0 and ηN, which change with differences in the compositions of the HMWGs.
Common buckwheat (Fagopyrum esculentum M.) belongs to the eudicot family Polygonaceae, Fagopyrum Mill, and its seeds have high nutritional value. The mechanism of seed development of common buckwheat remains unclear at the molecular level and no genes related to seed size have been identified. In this study, we performed genome-wide transcriptome sequencing and analysis using common buckwheat seeds at 5 days post anthesis (DPA) and 10 DPA from two cultivars (large-seeded and small-seeded). A total of 259,895 transcripts were assembled, resulting in 187,034 unigenes with average length of 1097 bp and N50 of 1538 bp. Based on gene expression profiles, 9127 differentially expressed genes (DEGs) were identified and analyzed in GO enrichment and KEGG analysis. In addition, genes related to seed size in the IKU pathway, ubiquitin–proteasome pathway, MAPK signaling pathway, TFs and phytohormones were identified and analyzed. AP2 and bZIP transcription factors, BR-signal and ABA were considered to be important regulators of seed size. This study provides a valuable genetic resource for future identification and functional analysis of candidate genes regulating seed size in common buckwheat and will be useful for improving seed yield in common buckwheat through molecular breeding in the future.
A core collection of eggplant (Solanum melongena L.) was developed based on a dataset of genome-wide 831 SNP and 50 SSR genotypes analyzed in 893 accessions of eggplant genetic resources collected in the NARO Genebank using the Core Hunter II program. The 893 accessions were collected worldwide, mainly Asia. Genetic variation and population structure among the 893 eggplant accessions were characterized. The genetic diversity of the Asian accessions, especially the South Asian and Southeast Asian accessions, forming the center of diversity in eggplant, was higher than that of the other regions. The resulting core collection, World Eggplant Core (WEC) collection consisted of 100 accessions basically collected from the high genetic diversity countries. Based on the results of the cluster and STRUCTURE analyses with SNP genotypes, the WEC collection was divided into four clusters (S1–S4). Each cluster corresponds to a geographical group as below, S1; the European, American and African countries, S2; the East Asian countries, S3; the Southeast Asian countries, S4; the South Asian and Southeast Asian countries. The genotype and phenotype data of the WEC collection are available from the VegMarks database (https://vegmarks.nivot.affrc.go.jp/resource/), and seed samples are available from the NARO Genebank (https://www.gene.affrc.go.jp/databases-core_collections.php).
Wheat processing quality is mainly correlated with high-molecular-weight glutenin subunits (HMW-GS) of grain endosperm. In bread wheat, the number of HMW-GS alleles are limited. However, wheat relative species possess numerous HMW-GS genes. In our previous study, a pair of novel HMW-GS 1Ux3.5+1Uy1.9 was characterized in Aegilops umbellulata. In this work, a novel wheat-Ae. umbellulata addition line, GN05, carrying a pair of 1U chromosome was developed and identified via cytogenetic analysis. Protein composition analysis indicated that GN05 carried HMW-GS of Ae. umbellulata. Accumulation of glutenin macropolymer (GMP) showed that GN05 had a much higher GMP content than the recurrent parent Chinese Spring. Rheological characteristics were analyzed by mixing test and the dough quality of GN05 was significantly improved compared to Chinese Spring. The results presented here may provide a valuable resource for the improvement of bread wheat quality.
Root system architecture (RSA) is one of the most important traits determining water and nutrient availability for plants. Modification of RSA is known to be a useful approach for improving root performance of crops. However, for conducting root phenotyping, there are few alternatives for the rapid collection of root samples from a constant soil volume. In this report, we propose a rapid root-sampling method, which uses a steel cylinder known as round monolith and backhoes to reduce the physical effort. The monolith was set on the ground surrounding individual rice plants and vertically driven back by a backhoe. Soil samples with 20 cm width and 25 cm depth were excavated by the monolith, from which root samples were then isolated. This backhoe-assisted monolith method requires at most five minutes to collect root samples from one plant. Using this method, we quantified the root traits of three rice lines, reported to form different types of root system such as shallow-, intermediate-, and deep-roots, using a root image analysis software. The data obtained through this method, which showed the same trend as previously reported, clearly demonstrated that this method is useful for quantitative evaluation of roots in the soil.
To breed new highly antioxidative common buckwheat cultivars, we selected individual plants from gamma ray-irradiated populations. Selection and propagation were repeated 4 or 5 times. This recurrent selection process resulted in many individuals with enhanced antioxidative activity. Among them, 2 individuals from the forth selection and 9 individuals from the fifth selection were developed into lines with increased antioxidative activities and diverse polyphenolic composition. From these lines, 2 new cultivars ‘Gamma no irodori’ and ‘Cobalt no chikara’ were developed. Furthermore, following the selection of individuals with high rutin contents, ‘Ruchiking’ was developed.
In Hokkaido, the northernmost region of Japan, soybean [Glycine max (L.) Merr.] crops are damaged by cold weather. Chilling temperatures negatively affect seed appearance by causing seed coat discoloration around the hilum region, which is called cold-induced discoloration (CD). An assay for CD tolerance using a phytotron was developed, and two quantitative trait loci (QTLs) associated with CD tolerance were identified. The major QTL was located in the proximal region of the I locus, and structural variation of this locus can serve as a useful DNA marker, called the Ic marker. To use this marker in breeding programs, the effects need to be assessed under field conditions because the Ic marker has been developed solely under phytotron conditions. The aim of this study was thus to assess the effect of the Ic marker under a cool field environment. We confirmed that the Ic allele was highly effective using 27 cultivars and breeding lines including a near-isogenic line grown in the field where severe cold-weather damage occurred. This allele had no negative influence on the agronomic traits in the near-isogenic line. Our results suggest that marker-assisted selection for the Ic allele is effective for improving CD tolerance in breeding programs.
Common cutworm (CCW) is a serious herbivorous insect pest of soybean. Previously, we conducted an antixenosis bioassay (measuring feeding preference) with CCW using recombinant inbred lines (RILs) derived from a cross between a wild soybean (Glycine soja) collected in Hiroshima prefecture (JP110755) and the leading cultivar, Fukuyutaka. The analysis revealed quantitative trait loci (QTLs) for antixenosis resistance, qRslx3 and qRslx4. In the present study we developed another RIL population using Fukuyutaka and a different G. soja, collected in Kumamoto prefecture (G406). An analysis revealed an antixenosis resistance QTL on chromosome 7, and the resistant allele of the QTL was derived from G406. The chromosomal position of the QTL was almost the same as that of CCW-2, a previously-reported antibiosis resistance QTL for CCW, detected in a F2 population derived from a cross between Fukuyutaka and a resistant cultivar Himeshirazu. These QTLs could be the same locus; however, G406 and Himeshirazu are likely to possess different alleles, because Himeshirazu allele exhibits no antixenosis effect. We expect that pyramiding of the resistance QTLs derived from G. soja will contribute to the development of CCW resistant cultivars.
The chrysanthemum (Chrysanthemum morifolium) is one of the most popular ornamental plants in the world. Genetic transformation is a promising tool for improving traits, editing genomes, and studying plant physiology. Promoters are vital components for efficient transformation, determining the level, location, and timing of transgene expression. The cauliflower mosaic virus (CaMV) 35S promoter is most frequently used in dicotyledonous plants but is less efficient in chrysanthemums than in tobacco or torenia plants. Previously, we used the parsley ubiquitin (PcUbi) promoter in chrysanthemums for the first time and analyzed its activity in transgenic calli. To expand the variety of constitutive promoters in chrysanthemums, we cloned the upstream region of the actin 2 (CmACT2) gene and compared its promoter activity with the 35S and PcUbi promoters in several organs, as well as its durability for long-term cultivation. The CmACT2 promoter has higher activity than the 35S promoter in calli but is less durable. The PcUbi promoter has the highest activity not only in calli but also in leaves, ray florets, and disk florets, and retains its activity after long-term cultivation. In conclusion, we have provided useful information and an additional type of promoter available for transgene expression in chrysanthemums.