Publications by authors named "Kenta Shirasawa"

97 Publications

Genome features of common vetch () in natural habitats.

Plant Direct 2021 Oct 7;5(10):e352. Epub 2021 Oct 7.

Kazusa DNA Research Institute Kisarazu Japan.

Wild plants are often tolerant to biotic and abiotic stresses in their natural environments, whereas domesticated plants such as crops frequently lack such resilience. This difference is thought to be due to the high levels of genome heterozygosity in wild plant populations and the low levels of heterozygosity in domesticated crop species. In this study, common vetch () was used as a model to examine this hypothesis. The common vetch genome (2n = 14) was estimated as 1.8 Gb in size. Genome sequencing produced a reference assembly that spanned 1.5 Gb, from which 31,146 genes were predicted. Using this sequence as a reference, 24,118 single nucleotide polymorphisms were discovered in 1243 plants from 12 natural common vetch populations in Japan. Common vetch genomes exhibited high heterozygosity at the population level, with lower levels of heterozygosity observed at specific genome regions. Such patterns of heterozygosity are thought to be essential for adaptation to different environments. The resources generated in this study will provide insights into de novo domestication of wild plants and agricultural enhancement.
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http://dx.doi.org/10.1002/pld3.352DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8496506PMC
October 2021

Genotyping-by-Sequencing Based Genetic Mapping Identified Major and Consistent Genomic Regions for Productivity and Quality Traits in Peanut.

Front Plant Sci 2021 23;12:668020. Epub 2021 Sep 23.

Department of Biotechnology, University of Agricultural Sciences, Dharwad, India.

With an objective of identifying the genomic regions for productivity and quality traits in peanut, a recombinant inbred line (RIL) population developed from an elite variety, TMV 2 and its ethyl methane sulfonate (EMS)-derived mutant was phenotyped over six seasons and genotyped with genotyping-by-sequencing (GBS), transposable element (AhTE) and simple sequence repeats (SSR) markers. The genetic map with 700 markers spanning 2,438.1 cM was employed for quantitative trait loci (QTL) analysis which identified a total of 47 main-effect QTLs for the productivity and oil quality traits with the phenotypic variance explained (PVE) of 10-52% over the seasons. A common QTL region (46.7-50.1 cM) on Ah02 was identified for the multiple traits, such as a number of pods per plant (NPPP), pod weight per plant (PWPP), shelling percentage (SP), and test weight (TW). Similarly, a QTL (7.1-18.0 cM) on Ah16 was identified for both SP and protein content (PC). Epistatic QTL (epiQTL) analysis revealed intra- and inter-chromosomal interactions for the main-effect QTLs and other genomic regions governing these productivity traits. The markers identified by a single marker analysis (SMA) mapped to the QTL regions for most of the traits. Among the five potential candidate genes identified for PC, SP and oil quality, two genes ( and ) were affected by transposition, and three genes (, and ) involved functional single nucleotide polymorphisms (SNPs). With major and consistent effects, the genomic regions, candidate genes, and the associated markers identified in this study would provide an opportunity for gene cloning and genomics-assisted breeding for increasing the productivity and enhancing the quality of peanut.
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http://dx.doi.org/10.3389/fpls.2021.668020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8495222PMC
September 2021

A chromosome-level genome sequence of Chrysanthemum seticuspe, a model species for hexaploid cultivated chrysanthemum.

Commun Biol 2021 Oct 7;4(1):1167. Epub 2021 Oct 7.

Graduate School of Integrated Sciences for Life, Hiroshima University, Higashi-Hiroshima, Hiroshima, Japan.

Chrysanthemums are one of the most industrially important cut flowers worldwide. However, their segmental allopolyploidy and self-incompatibility have prevented the application of genetic analysis and modern breeding strategies. We thus developed a model strain, Gojo-0 (Chrysanthemum seticuspe), which is a diploid and self-compatible pure line. Here, we present the 3.05 Gb chromosome-level reference genome sequence, which covered 97% of the C. seticuspe genome. The genome contained more than 80% interspersed repeats, of which retrotransposons accounted for 72%. We identified recent segmental duplication and retrotransposon expansion in C. seticuspe, contributing to arelatively large genome size. Furthermore, we identified a retrotransposon family, SbdRT, which was enriched in gene-dense genome regions and had experienced a very recent transposition burst. We also demonstrated that the chromosome-level genome sequence facilitates positional cloning in C. seticuspe. The genome sequence obtained here can greatly contribute as a reference for chrysanthemum in front-line breeding including genome editing.
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http://dx.doi.org/10.1038/s42003-021-02704-yDOI Listing
October 2021

Analysis of the Segregation Distortion of Genotypes Based on Whole-Genome Resequencing of Fig ( L.) Breeding Parents.

Front Plant Sci 2021 10;12:647599. Epub 2021 Aug 10.

Fukuoka Agriculture and Forestry Research Center Buzen Branch, Yukuhashi, Japan.

The common fig ( L.) has a gynodioecious breeding system, and its sex phenotype is an important trait for breeding because only female plant fruits are edible. During breeding to select for female plants, we analyzed the genotype, which is strongly associated with the sex phenotype. In 12 F populations derived from 13 cross combinations, the genotype segregation ratio was 1:1, whereas the M119-226 × H238-107 hybridization resulted in an extremely male-biased segregation ratio (178:7 = male:female). This finding suggests that the segregation distortion was caused by some genetic factor(s). A whole-genome resequencing of breeding parents (paternal and maternal lines) identified 9,061 high-impact SNPs in the parents. A genome-wide linkage analysis exploring the gene(s) responsible for the distortion revealed 194 high-impact SNPs specific to Caprifig6085 (i.e., seed parent ancestor) and 215 high-impact SNPs specific to H238-107 (i.e., pollen parent) in 201 annotated genes. A comparison between the annotated genes and the genes required for normal embryo or gametophyte development and function identified several candidate genes possibly responsible for the segregation distortion. This is the first report describing segregation distortion in .
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http://dx.doi.org/10.3389/fpls.2021.647599DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8384052PMC
August 2021

Chromosome-level genome assembly of Japanese chestnut (Castanea crenata Sieb. et Zucc.) reveals conserved chromosomal segments in woody rosids.

DNA Res 2021 Sep;28(5)

Kazusa DNA Research Institute, Chiba 292-0818, Japan.

Japanese chestnut (Castanea crenata Sieb. et Zucc.), unlike other Castanea species, is resistant to most diseases and wasps. However, genomic data of Japanese chestnut that could be used to determine its biotic stress resistance mechanisms have not been reported to date. In this study, we employed long-read sequencing and genetic mapping to generate genome sequences of Japanese chestnut at the chromosome level. Long reads (47.7 Gb; 71.6× genome coverage) were assembled into 781 contigs, with a total length of 721.2 Mb and a contig N50 length of 1.6 Mb. Genome sequences were anchored to the chestnut genetic map, comprising 14,973 single nucleotide polymorphisms (SNPs) and covering 1,807.8 cM map distance, to establish a chromosome-level genome assembly (683.8 Mb), with 69,980 potential protein-encoding genes and 425.5 Mb repetitive sequences. Furthermore, comparative genome structure analysis revealed that Japanese chestnut shares conserved chromosomal segments with woody plants, but not with herbaceous plants, of rosids. Overall, the genome sequence data of Japanese chestnut generated in this study is expected to enhance not only its genetics and genomics but also the evolutionary genomics of woody rosids.
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http://dx.doi.org/10.1093/dnares/dsab016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8435554PMC
September 2021

Genome-wide association study of individual sugar content in fruit of Japanese pear (Pyrus spp.).

BMC Plant Biol 2021 Aug 16;21(1):378. Epub 2021 Aug 16.

Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, 74 Kitainayazuma, Seika, Kyoto, 619-0244, Japan.

Background: Understanding mechanisms of sugar accumulation and composition is essential to determining fruit quality and maintaining a desirable balance of sugars in plant storage organs. The major sugars in mature Rosaceae fruits are sucrose, fructose, glucose, and sorbitol. Among these, sucrose and fructose have high sweetness, whereas glucose and sorbitol have low sweetness. Japanese pear has extensive variation in individual sugar contents in mature fruit. Increasing total sugar content and that of individual high-sweetness sugars is a major target of breeding programs. The objective of this study was to identify quantitative trait loci (QTLs) associated with fruit traits including individual sugar accumulation, to infer the candidate genes underlying the QTLs, and to assess the potential of genomic selection for breeding pear fruit traits.

Results: We evaluated 10 fruit traits and conducted genome-wide association studies (GWAS) for 106 cultivars and 17 breeding populations (1112 F1 individuals) using 3484 tag single-nucleotide polymorphisms (SNPs). By implementing a mixed linear model and a Bayesian multiple-QTL model in GWAS, 56 SNPs associated with fruit traits were identified. In particular, a SNP located close to acid invertase gene PPAIV3 on chromosome 7 and a newly identified SNP on chromosome 11 had quite large effects on accumulation of sucrose and glucose, respectively. We used 'Golden Delicious' doubled haploid 13 (GDDH13), an apple reference genome, to infer the candidate genes for the identified SNPs. In the region flanking the SNP on chromosome 11, there is a tandem repeat of early responsive to dehydration (ERD6)-like sugar transporter genes that might play a role in the phenotypes observed.

Conclusions: SNPs associated with individual sugar accumulation were newly identified at several loci, and candidate genes underlying QTLs were inferred using advanced apple genome information. The candidate genes for the QTLs are conserved across Pyrinae genomes, which will be useful for further fruit quality studies in Rosaceae. The accuracies of genomic selection for sucrose, fructose, and glucose with genomic best linear unbiased prediction (GBLUP) were relatively high (0.67-0.75), suggesting that it would be possible to select individuals having high-sweetness fruit with high sucrose and fructose contents and low glucose content.
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http://dx.doi.org/10.1186/s12870-021-03130-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8369641PMC
August 2021

DNA marker for resistance to in chrysanthemum ( Ramat.) "Southern Pegasus".

Breed Sci 2021 Apr 25;71(2):261-267. Epub 2021 Mar 25.

Institute of Fruit Tree and Tea Science, NARO, Tsukuba, Ibaraki 305-8605, Japan.

White rust caused by Henn. adversely affects chrysanthemum ( Ramat.) production. The breeding of resistant varieties is effective in controlling the disease. Here we aimed to develop DNA markers for the strong resistance to . We conducted a linkage analysis based on the genome-wide association study (GWAS) method. We employed a biparental population for the GWAS, wherein the single nucleotide polymorphism (SNP) allele frequency could be predicted. The population was derived from crosses between a strong resistant "Southern Pegasus" and a susceptible line. The GWAS used simplex and double-simplex SNP markers selected out of SNP candidates mined from ddRAD-Seq data of an F biparental population. These F individuals segregated in a 1:1 ratio of resistant to susceptible. Twenty-one simplex SNPs were significantly associated with resistance in "Southern Pegasus" and generated one linkage group. These results show the presence of a single resistance gene in "Southern Pegasus". We identified the nearest SNP marker located 2.2 cM from resistance locus and demonstrated this SNP marker-resistance link using an independent population. This is the first report of an effective DNA marker linked to a gene for resistance in chrysanthemum.
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http://dx.doi.org/10.1270/jsbbs.20063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8329880PMC
April 2021

Translation of continuous artificial selection on phenotype into genotype during rice breeding programs.

Breed Sci 2021 Apr 17;71(2):125-133. Epub 2021 Feb 17.

Kazusa DNA Research Institute, Kisarazu, Chiba 292-0818, Japan.

Understanding genetic diversity among local populations is a primary goal of modern crop breeding programs. Here, we demonstrated the genetic relationships of rice varieties in Hokkaido, Japan, one of the northern limits of rice cultivation around the world. Furthermore, artificial selection during rice breeding programs has been characterized using genome sequences. We utilized 8,565 single nucleotide polymorphisms and insertion/deletion markers distributed across the genome in genotype-by-sequencing for genetic diversity analyses. Phylogenetics, genetic population structure, and principal component analysis showed that a total of 110 varieties were classified into four distinct clusters according to different populations geographically and historically. Furthermore, the genome sequences of 19 rice varieties along with historic representations in Hokkaido, nucleotide diversity and values in each cluster revealed that artificial selection of elite phenotypes focused on chromosomal regions. These results clearly demonstrated the history of the selections on agronomic traits as genome sequences among current rice varieties from Hokkaido.
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http://dx.doi.org/10.1270/jsbbs.20089DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8329892PMC
April 2021

Chromosome-level genome assemblies of over 100 plant species.

Breed Sci 2021 Apr 6;71(2):117-124. Epub 2021 Apr 6.

ICAR-National Institute for Plant Biotechnology, Pusa, New Delhi 110012, India.

Genome sequence analysis in higher plants began with the whole-genome sequencing of . Owing to the great advances in sequencing technologies, also known as next-generation sequencing (NGS) technologies, genomes of more than 400 plant species have been sequenced to date. Long-read sequencing technologies, together with sequence scaffolding methods, have enabled the synthesis of chromosome-level genome sequence assemblies, which has further allowed comparative analysis of the structural features of multiple plant genomes, thus elucidating the evolutionary history of plants. However, the quality of the assembled chromosome-level sequences varies among plant species. In this review, we summarize the status of chromosome-level assemblies of 114 plant species, with genome sizes ranging from 125 Mb to 16.9 Gb. While the average genome coverage of the assembled sequences reached up to 89.1%, the average coverage of chromosome-level pseudomolecules was 73.3%. Thus, further improvements in sequencing technologies and scaffolding, and data analysis methods, are required to establish gap-free telomere-to-telomere genome sequence assemblies. With the forthcoming new technologies, we are going to enter into a new genomics era where pan-genomics and the >1,000 or >1 million genomes' project will be routine in higher plants.
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http://dx.doi.org/10.1270/jsbbs.20146DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8329882PMC
April 2021

Whole-genome sequencing and analysis of two azaleas, Rhododendron ripense and Rhododendron kiyosumense.

DNA Res 2021 Sep;28(5)

Kazusa DNA Research Institute, Kisarazu, Chiba 292-0818, Japan.

To enhance the genomics and genetics of azalea, the whole-genome sequences of two species of Rhododendron were determined and analysed in this study: Rhododendron ripense, the cytoplasmic donor and ancestral species of large-flowered and evergreen azalea cultivars; and Rhododendron kiyosumense, a native of Chiba prefecture (Japan) seldomly bred and cultivated. A chromosome-level genome sequence assembly of R. ripense was constructed by single-molecule real-time sequencing and genetic mapping, while the genome sequence of R. kiyosumense was assembled using the single-tube long fragment read sequencing technology. The R. ripense genome assembly contained 319 contigs (506.7 Mb; N50 length: 2.5 Mb) and was assigned to the genetic map to establish 13 pseudomolecule sequences. On the other hand, the genome of R. kiyosumense was assembled into 32,308 contigs (601.9 Mb; N50 length: 245.7 kb). A total of 34,606 genes were predicted in the R. ripense genome, while 35,785 flower and 48,041 leaf transcript isoforms were identified in R. kiyosumense through Iso-Seq analysis. Overall, the genome sequence information generated in this study enhances our understanding of genome evolution in the Ericales and reveals the phylogenetic relationship of closely related species. This information will also facilitate the development of phenotypically attractive azalea cultivars.
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http://dx.doi.org/10.1093/dnares/dsab010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8435550PMC
September 2021

Chromosome-scale genome assembly of the transformation-amenable common wheat cultivar 'Fielder'.

DNA Res 2021 Jun;28(3)

Kazusa DNA Research Institute, Kisarazu, 292-0818, Japan.

We have established a high-quality, chromosome-level genome assembly for the hexaploid common wheat cultivar 'Fielder', an American, soft, white, pastry-type wheat released in 1974 and known for its amenability to Agrobacterium tumefaciens-mediated transformation and genome editing. Accurate, long-read sequences were obtained using PacBio circular consensus sequencing with the HiFi approach. Sequence reads from 16 SMRT cells assembled using the hifiasm assembler produced assemblies with N50 greater than 20 Mb. We used the Omni-C chromosome conformation capture technique to order contigs into chromosome-level assemblies, resulting in 21 pseudomolecules with a cumulative size of 14.7 and 0.3 Gb of unanchored contigs. Mapping of published short reads from a transgenic wheat plant with an edited seed-dormancy gene, TaQsd1, identified four positions of transgene insertion into wheat chromosomes. Detection of guide RNA sequences in pseudomolecules provided candidates for off-target mutation induction. These results demonstrate the efficiency of chromosome-scale assembly using PacBio HiFi reads and their application in wheat genome-editing studies.
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http://dx.doi.org/10.1093/dnares/dsab008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8320877PMC
June 2021

Genomic Selection for F Hybrid Breeding in Strawberry ( × ).

Front Plant Sci 2021 4;12:645111. Epub 2021 Mar 4.

Department of Frontier Research and Development, Kazusa DNA Research Institute, Kisarazu, Japan.

Cultivated strawberry is the most widely consumed fruit crop in the world, and therefore, many breeding programs are underway to improve its agronomic traits such as fruit quality. Strawberry cultivars were vegetatively propagated through runners and carried a high risk of infection with viruses and insects. To solve this problem, the development of F hybrid seeds has been proposed as an alternative breeding strategy in strawberry. In this study, we conducted a potential assessment of genomic selection (GS) in strawberry F hybrid breeding. A total of 105 inbred lines were developed as candidate parents of strawberry F hybrids. In addition, 275 parental combinations were randomly selected from the 105 inbred lines and crossed to develop test F hybrids for GS model training. These populations were phenotyped for petiole length, leaf area, Brix, fruit hardness, and pericarp color. Whole-genome shotgun sequencing of the 105 inbred lines detected 20,811 single nucleotide polymorphism sites that were provided for subsequent GS analyses. In a GS model construction, inclusion of dominant effects showed a slight advantage in GS accuracy. In the across population prediction analysis, GS models using the inbred lines showed predictability for the test F hybrids and vice versa, except for Brix. Finally, the GS models were used for phenotype prediction of 5,460 possible F hybrids from 105 inbred lines to select F hybrids with high fruit hardness or high pericarp color. These F hybrids were developed and phenotyped to evaluate the efficacy of the GS. As expected, F hybrids that were predicted to have high fruit hardness or high pericarp color expressed higher observed phenotypic values than the F hybrids that were selected for other objectives. Through the analyses in this study, we demonstrated that GS can be applied for strawberry F hybrid breeding.
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http://dx.doi.org/10.3389/fpls.2021.645111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7969887PMC
March 2021

Genomic region associated with pod color variation in pea (Pisum sativum).

G3 (Bethesda) 2021 05;11(5)

Kazusa DNA Research Institute, Kisarazu, Chiba 292-0818, Japan.

Pea (Pisum sativum) was chosen as the research material by Gregor Mendel to discover the laws of inheritance. Out of seven traits studied by Mendel, genes controlling three traits including pod shape, pod color, and flower position have not been identified to date. With the aim of identifying the genomic region controlling pod color, we determined the genome sequence of a pea line with yellow pods. Genome sequence reads obtained using a Nanopore sequencing technology were assembled into 117,981 contigs (3.3 Gb), with an N50 value of 51.2 kb. A total of 531,242 potential protein-coding genes were predicted, of which 519,349 (2.8 Gb) were located within repetitive sequences (2.8 Gb). The assembled sequences were ordered using a reference as a guide to build pseudomolecules. Subsequent genetic and association analyses led to the identification of a genomic region that controls pea pod color. DNA sequences at this genomic location and transcriptome profiles of green and yellow pod lines were analyzed, and genes encoding 3' exoribonucleases were selected as potential candidates controlling pod color. The results presented in this study are expected to accelerate pan-genome studies in pea and facilitate the identification of the gene controlling one of the traits studied by Mendel.
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http://dx.doi.org/10.1093/g3journal/jkab081DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8104947PMC
May 2021

Chromosome-scale genome assembly of Japanese pear (Pyrus pyrifolia) variety 'Nijisseiki'.

DNA Res 2021 May;28(2)

Department of Frontier Research and Development, Kazusa DNA Research Institute, Chiba, Japan.

We analyzed the genome sequence of a Japanese pear (Pyrus pyrifolia) to facilitate its genetics and genomics as well as breeding programs, in which a variety 'Nijisseiki' with superior flesh texture has been used as a parent for most Japanese pear cultivars. De novo assembly of long sequence reads covered 136× of the Japanese pear genome and generated 503.9 Mb contigs consisting of 114 sequences with an N50 value of 7.6 Mb. Contigs were assigned to Japanese pear genetic maps to establish 17 chromosome-scale sequences. In total, 44,876 high-confidence protein-encoding genes were predicted, 84.3% of which were supported by predicted genes and transcriptome data from Japanese pear relatives. As expected, evidence of genome-wide duplication was observed, consistent with related species. This is the first chromosome-scale genome sequence analysis reported for Japanese pear, and this resource will support breeding programs and provide new insights into the physiology and evolutionary history of Japanese pear.
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http://dx.doi.org/10.1093/dnares/dsab001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8092371PMC
May 2021

De novo genome assembly of two tomato ancestors, Solanum pimpinellifolium and Solanum  lycopersicum var. cerasiforme, by long-read sequencing.

DNA Res 2021 Jan;28(1)

Tsukuba Plant Innovation Research Center, University of Tsukuba, Tsukuba, Ibaraki, 305-8572, Japan.

The ancestral tomato species are known to possess genes that are valuable for improving traits in breeding. Here, we aimed to construct high-quality de novo genome assemblies of Solanum pimpinellifolium 'LA1670' and S. lycopersicum var. cerasiforme 'LA1673', originating from Peru. The Pacific Biosciences (PacBio) long-read sequences with 110× and 104× coverages were assembled and polished to generate 244 and 202 contigs spanning 808.8 Mbp for 'LA1670' and 804.5 Mbp for 'LA1673', respectively. After chromosome-level scaffolding with reference guiding, 14 scaffold sequences corresponding to 12 tomato chromosomes and 2 unassigned sequences were constructed. High-quality genome assemblies were confirmed using the Benchmarking Universal Single-Copy Orthologs and long terminal repeat assembly index. The protein-coding sequences were then predicted, and their transcriptomes were confirmed. The de novo assembled genomes of S. pimpinellifolium and S. lycopersicum var. cerasiforme were predicted to have 71,945 and 75,230 protein-coding genes, including 29,629 and 29,185 non-redundant genes, respectively, as supported by the transcriptome analysis results. The chromosome-level genome assemblies coupled with transcriptome data sets of the two accessions would be valuable for gaining insights into tomato domestication and understanding genome-scale breeding.
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http://dx.doi.org/10.1093/dnares/dsaa029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7934570PMC
January 2021

Genome sequence of Hydrangea macrophylla and its application in analysis of the double flower phenotype.

DNA Res 2021 Jan;28(1)

Tochigi Prefectural Agricultural Experimental Station, Utsunomiya, Tochigi 320-0002, Japan.

Owing to its high ornamental value, the double flower phenotype of hydrangea (Hydrangea macrophylla) is one of its most important traits. In this study, genome sequence information was obtained to explore effective DNA markers and the causative genes for double flower production in hydrangea. Single-molecule real-time sequencing data followed by a Hi-C analysis were employed. Two haplotype-phased sequences were obtained from the heterozygous genome of hydrangea. One assembly consisted of 3,779 scaffolds (2.256 Gb in length and N50 of 1.5 Mb), the other also contained 3,779 scaffolds (2.227 Gb in length, and N50 of 1.4 Mb). A total of 36,930 genes were predicted in the sequences, of which 32,205 and 32,222 were found in each haplotype. A pair of 18 pseudomolecules was constructed along with a high-density single-nucleotide polymorphism (SNP) genetic linkage map. Using the genome sequence data, and two F2 populations, the SNPs linked to double flower loci (djo and dsu) were discovered. DNA markers linked to djo and dsu were developed, and these could distinguish the recessive double flower allele for each locus, respectively. The LEAFY gene is a very likely candidate as the causative gene for dsu, since frameshift was specifically observed in the double flower accession with dsu.
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http://dx.doi.org/10.1093/dnares/dsaa026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7934569PMC
January 2021

Four genetic loci control compact plant size with yellow pear-shaped fruit in ornamental tomato (Solanum lycopersicum L.).

Plant Genome 2020 07 11;13(2):e20017. Epub 2020 Apr 11.

Department of Frontier Research and Development, Kazusa DNA Research Institute, Kisarazu, Chiba, 292-0818, Japan.

Tomato is an attractive fruiting vegetable crop that can be used as an ornamental plant. Agronomical traits have been subjected to extensive genetic dissection to enhance vegetable breeding programs. By contrast, there are few genetic studies of ornamental traits for the development of ornamental tomato varieties. To investigate genetic loci linked to desired ornamental traits, we performed genetic analyses using an intraspecific mapping population that segregated for fruit color (yellow or red), fruit shape (round or pear), and plant height (high or compact). A genetic map was constructed with 965 single nucleotide polymorphisms (SNPs) and 33 simple sequence repeat markers. Subsequent linkage analysis using quantitative locus analysis and genome-wide association study detected four genetic loci for the three selected traits, all of which were located near the reported genes. We performed KASP-kompetitive allele-specific PCR-to develop SNP markers that were tightly linked to the four loci. Highly accurate genotyping data were obtained from the four SNPs across 187 F2 plants, which enabled us to select two lines with homozygous alleles for compact plant size and yellow pear-shaped fruits. These newly developed SNP markers and genetic strategies could be used to accelerate breeding programs for ornamental tomato plants.
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http://dx.doi.org/10.1002/tpg2.20017DOI Listing
July 2020

Genomic insight into the developmental history of southern highbush blueberry populations.

Heredity (Edinb) 2021 01 1;126(1):194-205. Epub 2020 Sep 1.

Graduate School of Agriculture, Kyoto University, Sakyo-Ku, Kyoto, 606-8502, Japan.

Interspecific hybridization is a common breeding approach for introducing novel traits and genetic diversity to breeding populations. Southern highbush blueberry (SHB) is a blueberry cultivar group that has been intensively bred over the last 60 years. Specifically, it was developed by multiple interspecific crosses between northern highbush blueberry [NHB, Vaccinium corymbosum L. (2n = 4x = 48)] and low-chill Vaccinium species to expand the geographic limits of highbush blueberry production. In this study, we genotyped polyploid blueberries, including 105 SHB, 17 NHB, and 10 rabbiteye blueberry (RE) (Vaccinium virgatum Aiton), from the accessions planted at Poplarville, Mississippi, and accessions distributed in Japan, based on the double-digest restriction site-associated DNA sequencing. The genome-wide SNP data clearly indicated that RE cultivars were genetically distinct from SHB and NHB cultivars, whereas NHB and SHB were genetically indistinguishable. The population structure results appeared to reflect the differences in the allele selection strategies that breeders used for developing germplasm adapted to local climates. The genotype data implied that there are no or very few genomic segments that were commonly introgressed from low-chill Vaccinium species to the SHB genome. Principal component analysis-based outlier detection analysis found a few loci associated with a variable that could partially differentiate NHB and SHB. These SNP loci were detected in Mb-scale haplotype blocks and may be close to the functional genes related to SHB development. Collectively, the data generated in this study suggest a polygenic adaptation of SHB to the southern climate, and may be relevant for future population-scale genome-wide analyses of blueberry.
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http://dx.doi.org/10.1038/s41437-020-00362-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7853091PMC
January 2021

Development of High-Density Genetic Linkage Maps and Identification of Loci for Chestnut Gall Wasp Resistance in spp.

Plants (Basel) 2020 Aug 18;9(8). Epub 2020 Aug 18.

Dipartimento di Scienze Agrarie, Forestali e Alimentari-DISAFA, Università degli Studi di Torino, Largo Paolo Braccini 2, Grugliasco, 10095 Torino, Italy.

is an important multipurpose species in Europe for nut and timber production as well as for its role in the landscape and in the forest ecosystem. This species has low tolerance to chestnut gall wasp ( Yasumatsu), which is a pest that was accidentally introduced into Europe in early 2000 and devastated forest and orchard trees. Resistance to the gall wasp was found in the hybrid cultivar 'Bouche de Bétizac' ( × ) and studied by developing genetic linkage maps using a population derived from a cross between 'Bouche de Bétizac' and the susceptible cultivar 'Madonna' (). The high-density genetic maps were constructed using double-digest restriction site-associated DNA-seq and simple sequence repeat markers. The map of 'Bouche de Bétizac' consisted of 1459 loci and spanned 809.6 cM; the map of 'Madonna' consisted of 1089 loci and spanned 753.3 cM. In both maps, 12 linkage groups were identified. A single major QTL was recognized on the 'Bouche de Bétizac' map, explaining up to 67-69% of the phenotypic variance of the resistance trait (). The quantitative trait loci (QTL) region included 11 scaffolds and two candidate genes putatively involved in the resistance response were identified. This study will contribute to breeding programs and to the study of genes.
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http://dx.doi.org/10.3390/plants9081048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7465717PMC
August 2020

Genome-wide study on the polysomic genetic factors conferring plasticity of flower sexuality in hexaploid persimmon.

DNA Res 2020 Jun;27(3)

Graduate School of Environmental and Life Science, Okayama University, Okayama 700-8530, Japan.

Sexuality is one of the fundamental mechanisms that work towards maintaining genetic diversity within a species. In diploid persimmons (Diospyros spp.), separated sexuality, the presence of separate male and female individuals (dioecy), is controlled by the Y chromosome-encoded small-RNA gene, OGI. On the other hand, sexuality in hexaploid Oriental persimmon (Diospyros kaki) is more plastic, with OGI-bearing genetically male individuals, able to produce both male and female flowers (monoecy). This is thought to be linked to the partial inactivation of OGI by a retrotransposon insertion, resulting in DNA methylation of the OGI promoter region. To identify the genetic factors regulating branch sexual conversion, genome-wide correlation/association analyses were conducted using ddRAD-Seq data from an F1 segregating population, and using both quantitative and diploidized genotypes, respectively. We found that allelic ratio at the Y-chromosomal region, including OGI, was correlated with male conversion based on quantitative genotypes, suggesting that OGI can be activated in cis in a dosage-dependent manner. Genome-wide association analysis based on diploidized genotypes, normalized for the effect of OGI allele dosage, detected three fundamental loci associated with male conversion. These loci underlie candidate genes, which could potentially act epigenetically for the activation of OGI expression.
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http://dx.doi.org/10.1093/dnares/dsaa012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7406971PMC
June 2020

QTL analysis and GWAS of agronomic traits in sweetpotato ( L.) using genome wide SNPs.

Breed Sci 2020 Jun 19;70(3):283-291. Epub 2020 May 19.

Kyushu Okinawa Agricultural Research Center, NARO, 6651-2 Yokoichi-cho, Miyakonojo, Miyazaki 885-0091, Japan.

While sweetpotato ( L.) improvement has generally been done by field-based selection, molecular genetic studies on traits of interest, i.e., molecular markers are needed for enhancing the breeding program of this world's 7 most important crop, as such markers facilitate marker-assisted selection. Here, we performed a combined approach of QTLs analyses and GWAS of storage root β-carotene content (BC), dry-matter (DM) and starch content (SC) using the genetic linkage maps constructed with 5,952 and 5,640 SNPs obtained from F progenies between cultivars 'J-Red' and 'Choshu'. BC was negatively correlated with DM (r = -0.45) and SC (r = -0.51), while DM was positively correlated with SC (r = 0.94). In both parental maps, a total of five, two and five QTL regions on linkage groups 7 and 8 were associated with BC, DM and SC, respectively. In GWAS of BC, one strong signal ( = 1.04 × 10) was observed on linkage group 8, which co-located with one of the above QTL regions. The SNPs markers found here, particularly for β-carotene, would be useful base resources for future marker-assisted selection program with this trait.
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http://dx.doi.org/10.1270/jsbbs.19099DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7372034PMC
June 2020

CURLED LATER1 encoding the largest subunit of the Elongator complex has a unique role in leaf development and meristem function in rice.

Plant J 2020 10 31;104(2):351-364. Epub 2020 Jul 31.

School of Science, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo, 113-8654, Japan.

The Elongator complex, which is conserved in eukaryotes, has multiple roles in diverse organisms. In Arabidopsis thaliana, Elongator is shown to be involved in development, hormone action and environmental responses. However, except for Arabidopsis, our knowledge of its function is poor in plants. In this study, we initially carried out a genetic analysis to characterize a rice mutant with narrow and curled leaves, termed curled later1 (cur1). The cur1 mutant displayed a heteroblastic change, whereby the mutant leaf phenotype appeared specifically at a later adult phase of vegetative development. The shoot apical meristem (SAM) was small and the leaf initiation rate was low, suggesting that the activity of the SAM seemed to be partially reduced in cur1. We then revealed that CUR1 encodes a yeast ELP1-like protein, the largest subunit of Elongator. Furthermore, disruption of OsELP3 encoding the catalytic subunit of Elongator resulted in phenotypes similar to those of cur1, including the timing of the appearance of mutant phenotypes. Thus, Elongator activity seems to be specifically required for leaf development at the late vegetative phase. Transcriptome analysis showed that genes involved in protein quality control were highly upregulated in the cur1 shoot apex at the later vegetative phase, suggesting the restoration of impaired proteins probably produced by partial defects in translational control due to the loss of function of Elongator. The differences in the mutant phenotype and gene expression profile between CUR1 and its Arabidopsis ortholog suggest that Elongator has evolved to play a unique role in rice development.
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http://dx.doi.org/10.1111/tpj.14925DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7689840PMC
October 2020

Root-knot nematode genetic diversity associated with host compatibility to sweetpotato cultivars.

Mol Plant Pathol 2020 08 17;21(8):1088-1098. Epub 2020 Jun 17.

Faculty of Agriculture, Ryukoku University, Otsu, Shiga, Japan.

Plant parasitic root-knot nematodes (RKN) such as Meloidogyne incognita cause significant crop losses worldwide. Although RKN are polyphagous, with wide host ranges, races with differing host compatibilities have evolved. Associations between genotype and infection phenotype in M. incognita have not yet been discovered. In this study, 48 M. incognita isolates were collected from geographically diverse fields in Japan and their genomes sequenced. The isolates exhibited various infection compatibilities to five sweetpotato (SP) cultivars and were assigned to SP races. Genome-wide association analysis identified 743 SNPs affecting gene coding sequences, a large number of which (575) were located on a single 1 Mb region. To examine how this polymorphic region evolved, nucleotide diversity (Pi) was scanned at the whole genome scale. The SNP-rich 1 Mb region exhibited high Pi values and was clearly associated with the SP races. SP1 and 2 races showed high Pi values in this region whereas the Pi values of SP3, 4, and 6 were low. Principal component analysis of isolates from this study and globally collected isolates showed selective divergence in this 1 Mb region. Our results suggest for the first time that the host could be a key determining factor stimulating the genomic divergence of M. incognita.
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http://dx.doi.org/10.1111/mpp.12961DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7368124PMC
August 2020

Genetic Mapping in Autohexaploid Sweet Potato with Low-Coverage NGS-Based Genotyping Data.

G3 (Bethesda) 2020 08 5;10(8):2661-2670. Epub 2020 Aug 5.

Kazusa DNA Research Institute, Japan.

Next-generation sequencing (NGS)-based genotyping methods can generate numerous genetic markers in a single experiment and have contributed to plant genetic mapping. However, for high precision genetic analysis, the complicated genetic segregation mode in polyploid organisms requires high-coverage NGS data and elaborate analytical algorithms. In the present study, we propose a simple strategy for the genetic mapping of polyploids using low-coverage NGS data. The validity of the strategy was investigated using simulated data. Previous studies indicated that accurate allele dosage estimation from low-coverage NGS data (read depth < 40) is difficult. Therefore, we used allele dosage probabilities calculated from read counts in association analyses to detect loci associated with phenotypic variations. The allele dosage probabilities showed significant detection power, although higher allele dosage estimation accuracy resulted in higher detection power. On the contrary, differences in the segregation patterns between the marker and causal genes resulted in a drastic decrease in detection power even if the marker and casual genes were in complete linkage and the allele dosage estimation was accurate. These results indicated that the use of a larger number of markers is advantageous, even if the accuracy of allele dosage estimation is low. Finally, we applied the strategy for the genetic mapping of autohexaploid sweet potato () populations to detect loci associated with agronomic traits. Our strategy could constitute a cost-effective approach for preliminary experiments done performed to large-scale studies.
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http://dx.doi.org/10.1534/g3.120.401433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7407471PMC
August 2020

Genome sequence and analysis of a Japanese radish (Raphanus sativus) cultivar named 'Sakurajima Daikon' possessing giant root.

DNA Res 2020 Apr;27(2)

Kazusa DNA Research Institute, Kisarazu, Chiba 292-0818, Japan.

Aim: The complex genome of a Japanese radish (Raphanus sativus) cultivar named 'Okute-Sakurajima' with an extremely large edible round root was analysed to explore its genomic characteristics.

Methods And Results: Single-molecule real-time technology was used to obtain long sequence reads to cover 60× of the genome. De novo assembly generated 504.5 Mb contigs consisting of 1,437 sequences with the N50 value of 1.2 Mb and included 94.1% of the core eukaryotic genes. Nine pseudomolecules, comprising 69.3% of the assembled contigs, were generated along with a high-density SNP genetic map. The sequence data thus established revealed the presence of structural variations and rearrangements in the Brassicaceae genomes.

Conclusion And Perspective: A total of 89,915 genes were identified in the 'Okute-Sakurajima' genome, 30,033 of which were newly found in this study. The genome information reported here will not only contribute to the establishment of a new resource for the radish genomics but also provide insights into the molecular mechanisms underlying formation of the giant root.
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http://dx.doi.org/10.1093/dnares/dsaa010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7334891PMC
April 2020

The persimmon genome reveals clues to the evolution of a lineage-specific sex determination system in plants.

PLoS Genet 2020 02 18;16(2):e1008566. Epub 2020 Feb 18.

Genome Center and Department of Plant Biology, University of California Davis, Davis, California, United States of America.

Most angiosperms bear hermaphroditic flowers, but a few species have evolved outcrossing strategies, such as dioecy, the presence of separate male and female individuals. We previously investigated the mechanisms underlying dioecy in diploid persimmon (D. lotus) and found that male flowers are specified by repression of the autosomal gene MeGI by its paralog, the Y-encoded pseudo-gene OGI. This mechanism is thought to be lineage-specific, but its evolutionary path remains unknown. Here, we developed a full draft of the diploid persimmon genome (D. lotus), which revealed a lineage-specific whole-genome duplication event and provided information on the architecture of the Y chromosome. We also identified three paralogs, MeGI, OGI and newly identified Sister of MeGI (SiMeGI). Evolutionary analysis suggested that MeGI underwent adaptive evolution after the whole-genome duplication event. Transformation of tobacco plants with MeGI and SiMeGI revealed that MeGI specifically acquired a new function as a repressor of male organ development, while SiMeGI presumably maintained the original function. Later, a segmental duplication event spawned MeGI's regulator OGI on the Y-chromosome, completing the path leading to dioecy, and probably initiating the formation of the Y-chromosome. These findings exemplify how duplication events can provide flexible genetic material available to help respond to varying environments and provide interesting parallels for our understanding of the mechanisms underlying the transition into dieocy in plants.
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http://dx.doi.org/10.1371/journal.pgen.1008566DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7048303PMC
February 2020

Identification of genome-wide single-nucleotide polymorphisms among geographically diverse radish accessions.

DNA Res 2020 Feb;27(1)

Graduate School of Agricultural Science, Tohoku University, Aoba-ku, Sendai 980-8572, Japan.

Radish (Raphanus sativus L.) is cultivated around the world as a vegetable crop and exhibits diverse morphological and physiological features. DNA polymorphisms are responsible for differences in traits among cultivars. In this study, we determined genome-wide single-nucleotide polymorphisms (SNPs) among geographically diverse radish accessions using the double-digest restriction site-associated DNA sequencing (ddRAD-Seq) method. A total of 52,559 SNPs was identified in a collection of over 500 radish accessions (cultivated and wild) from East Asia, South and Southeast Asia, and the Occident and Near East. In addition, 2,624 SNP sites without missing data (referred to as common SNP sites) were identified among 510 accessions. Genetic diversity analyses, based on the common SNP sites, divided the cultivated radish accessions into four main groups, each derived from four geographical areas (Japan, East Asia, South and Southeast Asia, and the Occident and Near East). Furthermore, we discuss the origin of cultivated radish and its migration from the West to East Asia. SNP data generated in this work will facilitate further genetic studies on the radish breeding and production of DNA markers.
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http://dx.doi.org/10.1093/dnares/dsaa001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7315352PMC
February 2020

The Ficus erecta genome aids Ceratocystis canker resistance breeding in common fig (F. carica).

Plant J 2020 06 24;102(6):1313-1322. Epub 2020 Feb 24.

Kazusa DNA Research Institute, Kisarazu, Japan.

Ficus erecta, a wild relative of the common fig (F. carica), is a donor of Ceratocystis canker resistance in fig breeding programmes. Interspecific hybridization followed by recurrent backcrossing is an effective method to transfer the resistance trait from wild to cultivated fig. However, this process is time consuming and labour intensive for trees, especially for gynodioecious plants such as fig. In this study, genome resources were developed for F. erecta to facilitate fig breeding programmes. The genome sequence of F. erecta was determined using single-molecule real-time sequencing technology. The resultant assembly spanned 331.6 Mb with 538 contigs and an N50 length of 1.9 Mb, from which 51 806 high-confidence genes were predicted. Pseudomolecule sequences corresponding to the chromosomes of F. erecta were established with a genetic map based on single nucleotide polymorphisms from double-digest restriction-site-associated DNA sequencing. Subsequent linkage analysis and whole-genome resequencing identified a candidate gene for the Ceratocystis canker resistance trait. Genome-wide genotyping analysis enabled the selection of female lines that possessed resistance and effective elimination of the donor genome from the progeny. The genome resources provided in this study will accelerate and enhance disease-resistance breeding programmes in fig.
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http://dx.doi.org/10.1111/tpj.14703DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7317799PMC
June 2020

Analysis of the lineage of Phytophthora infestans isolates using mating type assay, traditional markers, and next generation sequencing technologies.

PLoS One 2020 21;15(1):e0221604. Epub 2020 Jan 21.

Department of Frontier Research and Development, Kazusa DNA Research Institute, Chiba, Japan.

Phytophthora infestans (Mont.) de Bary, a hemibiotrophic oomycete, has caused severe epidemics of late blight in tomato and potato crops around the world since the Irish Potato Famine in the 1840s. Breeding of late blight resistant cultivars is one of the most effective strategies to overcome this disruptive disease. However, P. infestans is able to break down host resistance and acquire resistance to various fungicides, possibly because of the existence of high genetic variability among P. infestans isolates via sexual and asexual reproduction. Therefore, to manage this disease, it is important to understand the genetic divergence of P. infestans isolates. In this study, we analyzed the genomes of P. infestans isolates collected from Egypt and Japan using various molecular approaches including the mating type assay and genotyping simple sequence repeats, mitochondria DNA, and effector genes. We also analyzed genome-wide single nucleotide polymorphisms using double-digest restriction-site associated DNA sequencing and whole genome resequencing (WGRS). The isolates were classified adequately using high-resolution genome-wide approaches. Moreover, these analyses revealed new clusters of P. infestans isolates in the Egyptian population. Monitoring the genetic divergence of P. infestans isolates as well as breeding of resistant cultivars would facilitate the elimination of the late blight disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0221604PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6974037PMC
April 2020
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