Different control of resistance to two Colletotrichum orbiculare pathogenic races 0 and 1 in cucumber (Cucumis sativus L.).

KEY MESSAGE: Quantitative trait locus analysis identified independent novel loci in cucumbers responsible for resistance to races 0 and 1 of the anthracnose fungal pathogen Colletotrichum orbiculare. Cucumbers have been reported to be vulnerable to Colletotrichum orbiculare, causing anthracnose disease with significant yield loss under favorable conditions. The deployment of a single recessive Cssgr gene in cucumber breeding for anthracnose resistance was effective until a recent report on high-virulent strains infecting cucumbers in Japan conquering the resistance. QTL mapping was conducted to identify the resistance loci in the cucumber accession Ban Kyuri (G100) against C. orbiculare strains 104-T and CcM-1 of pathogenic races 0 and 1, respectively. A single dominant locus An5 was detected in the disease resistance hotspot on chromosome 5 for resistance to 104-T. Resistance to CcM-1 was governed by three loci with additive effects located on chromosomes 2 (An2) and 1 (An1.1 and An1.2). Molecular markers were developed based on variant calling between the corresponding QTL regions in the de novo assembly of the G100 genome and the publicly available cucumber genomes. Multiple backcrossed populations were deployed to fine-map An5 locus and narrow the region to approximately 222 kbp. Accumulation of An2 and An1.1 alleles displayed an adequate resistance to CcM-1 strain. This study provides functional molecular markers for pyramiding resistance loci that confer sufficient resistance against anthracnose in cucumbers.

Assessing the Potential for Genome-Assisted Breeding in Red Perilla Using Quantitative Trait Locus Analysis and Genomic Prediction.

Red perilla is an important medicinal plant used in Kampo medicine. The development of elite varieties of this species is urgently required. Medicinal compounds are generally considered target traits in medicinal plant breeding; however, selection based on compound phenotypes (i.e., conventional selection) is expensive and time consuming. Here, we propose genomic selection (GS) and marker-assisted selection (MAS), which use marker information for selection, as suitable selection methods for medicinal plants, and we evaluate the effectiveness of these methods in perilla breeding. Three breeding populations generated from crosses between one red and three green perilla genotypes were used to elucidate the genetic mechanisms underlying the production of major medicinal compounds using quantitative trait locus analysis and evaluating the accuracy of genomic prediction (GP). We found that GP had a sufficiently high accuracy for all traits, confirming that GS is an effective method for perilla breeding. Moreover, the three populations showed varying degrees of segregation, suggesting that using these populations in breeding may simultaneously enhance multiple target traits. This study contributes to research on the genetic mechanisms of the major medicinal compounds of red perilla, as well as the breeding efficiency of this medicinal plant.

Telomere-to-telomere genome assembly of an allotetraploid pernicious weed, Echinochloa phyllopogon.

Echinochloa phyllopogon is an allotetraploid pernicious weed species found in rice fields worldwide that often exhibit resistance to multiple herbicides. An accurate genome sequence is essential to comprehensively understand the genetic basis underlying the traits of this species. Here, the telomere-to-telomere genome sequence of E. phyllopogon was presented. Eighteen chromosome sequences spanning 1.0 Gb were constructed using the PacBio highly fidelity long technology. Of the 18 chromosomes, 12 sequences were entirely assembled into telomere-to-telomere and gap-free contigs, whereas the remaining six sequences were constructed at the chromosomal level with only eight gaps. The sequences were assigned to the A and B genome with total lengths of 453 and 520 Mb, respectively. Repetitive sequences occupied 42.93% of the A genome and 48.47% of the B genome, although 32,337, and 30,889 high-confidence genes were predicted in the A and B genomes, respectively. This suggested that genome extensions and gene disruptions caused by repeated sequence accumulation often occur in the B genome before polyploidization to establish a tetraploid genome. The highly accurate and comprehensive genome sequence could be a milestone in understanding the molecular mechanisms of the pernicious traits and in developing effective weed control strategies to avoid yield loss in rice production.

Whole-genome sequence analysis of mutations in rice plants regenerated from zygotes, mature embryos, and immature embryos.

Somaclonal variation was studied by whole-genome sequencing in rice plants (Oryza sativa L., 'Nipponbare') regenerated from the zygotes, mature embryos, and immature embryos of a single mother plant. The mother plant and its seed-propagated progeny were also sequenced. A total of 338 variants of the mother plant sequence were detected in the progeny, and mean values ranged from 9.0 of the seed-propagated plants to 37.4 of regenerants from mature embryos. The natural mutation rate of 1.2 × 10-8 calculated using the variants in the seed-propagated plants was consistent with the values reported previously. The ratio of single nucleotide variants (SNVs) among the variants in the seed-propagated plants was 91.1%, which is higher than 56.1% previously reported, and not significantly different from those in the regenerants. Overall, the ratio of transitions to transversions of SNVs was lower in the regenerants as shown previously. Plants regenerated from mature embryos had significantly more variants than different progeny types. Therefore, using zygotes and immature embryos can reduce somaclonal variation during the genetic manipulation of rice.

A Novel Method for Quantifying Plant Morphological Characteristics Using Normal Vectors and Local Curvature Data via 3D Modelling-A Case Study in Leaf Lettuce.

Three-dimensional measurement is a high-throughput method that can record a large amount of information. Three-dimensional modelling of plants has the possibility to not only automate dimensional measurement, but to also enable visual assessment to be quantified, eliminating ambiguity in human judgment. In this study, we have developed new methods that could be used for the morphological analysis of plants from the information contained in 3D data. Specifically, we investigated characteristics that can be measured by scale (dimension) and/or visual assessment by humans. The latter is particularly novel in this paper. The characteristics that can be measured on a scale-related dimension were tested based on the bounding box, convex hull, column solid, and voxel. Furthermore, for characteristics that can be evaluated by visual assessment, we propose a new method using normal vectors and local curvature (LC) data. For these examinations, we used our highly accurate all-around 3D plant modelling system. The coefficient of determination between manual measurements and the scale-related methods were all above 0.9. Furthermore, the differences in LC calculated from the normal vector data allowed us to visualise and quantify the concavity and convexity of leaves. This technique revealed that there were differences in the time point at which leaf blistering began to develop among the varieties. The precise 3D model made it possible to perform quantitative measurements of lettuce size and morphological characteristics. In addition, the newly proposed LC-based analysis method made it possible to quantify the characteristics that rely on visual assessment. This research paper was able to demonstrate the following possibilities as outcomes: (1) the automation of conventional manual measurements, and (2) the elimination of variability caused by human subjectivity, thereby rendering evaluations by skilled experts unnecessary.

Plant GARDEN: a portal website for cross-searching between different types of genomic and genetic resources in a wide variety of plant species.

BACKGROUND: Plant genome information is fundamental to plant research and development. Along with the increase in the number of published plant genomes, there is a need for an efficient system to retrieve various kinds of genome-related information from many plant species across plant kingdoms. Various plant databases have been developed, but no public database covers both genomic and genetic resources over a wide range of plant species. MAIN BODY: We have developed a plant genome portal site, Plant GARDEN (Genome And Resource Database Entry: https://plantgarden.jp/en/index ), to provide diverse information related to plant genomics and genetics in divergent plant species. Elasticsearch is used as a search engine, and cross-keyword search across species is available. Web-based user interfaces (WUI) for PCs and tablet computers were independently developed to make data searches more convenient. Several types of data are stored in Plant GARDEN: reference genomes, gene sequences, PCR-based DNA markers, trait-linked DNA markers identified in genetic studies, SNPs, and in/dels on publicly available sequence read archives (SRAs). The data registered in Plant GARDEN as of March 2023 included 304 assembled genome sequences, 11,331,614 gene sequences, 419,132 DNA markers, 8,225 QTLs, and 5,934 SNP lists (gvcf files). In addition, we have re-annotated all the genes registered in Plant GARDEN by using a functional annotation tool, Hayai-Annotation, to compare the orthologous relationships among genes. CONCLUSION: The aim of Plant GARDEN is to provide plant genome information for use in the fields of plant science as well as for plant-based industries, education, and other relevant areas. Therefore, we have designed a WUI that allows a diverse range of users to access such information in an easy-to-understand manner. Plant GARDEN will eventually include a wide range of plant species for which genome sequences are assembled, and thus the number of plant species in the database will continue to expand. We anticipate that Plant GARDEN will promote the understanding of genomes and gene diversity by facilitating comparisons of the registered sequences.

Polyploid GWAS reveals the basis of molecular marker development for complex breeding traits including starch content in the storage roots of sweet potato.

Given the importance of prioritizing genome-based breeding of sweet potato to enable the promotion of food and nutritional security for future human societies, here, we aimed to dissect the genetic basis of storage root starch content (SC) when associated with a complex set of breeding traits including dry matter (DM) rate, storage root fresh weight (SRFW), and anthocyanin (AN) content in a mapping population containing purple-fleshed sweet potato. A polyploid genome-wide association study (GWAS) was extensively exploited using 90,222 single-nucleotide polymorphisms (SNPs) obtained from a bi-parental 204 F1 population between 'Konaishin' (having high SC but no AN) and 'Akemurasaki' (having high AN content but moderate SC). Through the comparison of polyploid GWAS on the whole set of the 204 F1, 93 high-AN-containing F1, and 111 low-AN-containing F1 populations, a total of two (consists of six SNPs), two (14 SNPs), four (eight SNPs), and nine (214 SNPs) significantly associated signals were identified for the variations of SC, DM, SRFW, and the relative AN content, respectively. Of them, a novel signal associated with SC, which was most consistent in 2019 and 2020 in both the 204 F1 and 111 low-AN-containing F1 populations, was identified in homologous group 15. The five SNP markers associated with homologous group 15 could affect SC improvement with a degree of positive effect (~4.33) and screen high-starch-containing lines with higher efficiency (~68%). In a database search of 62 genes involved in starch metabolism, five genes including enzyme genes granule-bound starch synthase I (IbGBSSI), α-amylase 1D, α-amylase 1E, and α-amylase 3, and one transporter gene ATP/ADP-transporter were located on homologous group 15. In an extensive qRT-PCR of these genes using the storage roots harvested at 2, 3, and 4 months after field transplantation in 2022, IbGBSSI, which encodes the starch synthase isozyme that catalyzes the biosynthesis of amylose molecule, was most consistently elevated during starch accumulation in sweet potato. These results would enhance our understanding of the underlying genetic basis of a complex set of breeding traits in the starchy roots of sweet potato, and the molecular information, particularly for SC, would be a potential platform for molecular marker development for this trait.

Identification of novel sex determination loci in Japanese weedy melon.

KEY MESSAGE: Japanese weedy melon exhibits unique sex expression with interactions between previously reported sex determination genes and two novel loci. Sex expression contributes to fruit quality and yield in the Cucurbitaceae. In melon, orchestrated regulation by sex determination genes explains the mechanism of sex expression, resulting in a great variety of sexual morphologies. In this study, we examined the Japanese weedy melon UT1, which does not follow the reported model of sex expression. We conducted QTL analysis using F2 plants for flower sex on the main stem and the lateral branch and mapped "occurrence of pistil-bearing flower on the main stem" locus on Chr. 3 (Opbf3.1) and "type of pistil-bearing flower" (female or bisexual) loci on Chr. 2 (tpbf2.1) and Chr. 8 (tpbf8.1). The Opbf3.1 included the known sex determination gene CmACS11. Sequence comparison of CmACS11 between parental lines revealed three nonsynonymous SNPs. A CAPS marker developed from one of the SNPs was closely linked to the occurrence of pistil-bearing flowers on the main stem in two F2 populations with different genetic backgrounds. The UT1 allele on Opbf3.1 was dominant in F1 lines from crosses between UT1 and diverse cultivars and breeding lines. This study suggests that Opbf3.1 and tpbf8.1 may promote the development of pistil and stamen primordia by inhibiting CmWIP1 and CmACS-7 functions, respectively, making the UT1 plants hermaphrodite. The results of this study provide new insights into the molecular mechanisms of sex determination in melons and considerations for the application of femaleness in melon breeding.

Telomere-to-telomere genome assembly of matsutake (Tricholoma matsutake).

Here, we report the first telomere-to-telomere genome assembly of matsutake (Tricholoma matsutake), which consists of 13 sequences (spanning 161.0 Mb) and a 76 kb circular mitochondrial genome. All the 13 sequences were supported with telomeric repeats at the ends. GC-rich regions are located at the middle of the sequences and are enriched with long interspersed nuclear elements (LINEs). Repetitive sequences including long-terminal repeats (LTRs) and LINEs occupy 71.6% of the genome. A total of 21,887 potential protein-coding genes were predicted. The genomic data reported in this study served not only matsutake gene sequences but also genome structures and intergenic sequences. The information gained would be a great reference for exploring the genetics, genomics, and evolutionary study of matsutake in the future, and ultimately facilitate the conservation of this vulnerable genetic resource.

Power Up Plant Genetic Research with Genomic Data.

The official debut of the reference genome of Arabidopsis thaliana in 2000 [...].

An improved reference genome for Trifolium subterraneum L. provides insight into molecular diversity and intra-specific phylogeny.

Subterranean clover (Trifolium subterraneum L., Ts) is a geocarpic, self-fertile annual forage legume with a compact diploid genome (n = x = 8, 544 Mb/1C). Its resilience and climate adaptivity have made it an economically important species in Mediterranean and temperate zones. Using the cultivar Daliak, we generated higher resolution sequence data, created a new genome assembly TSUd_3.0, and conducted molecular diversity analysis for copy number variant (CNV) and single-nucleotide polymorphism (SNP) among 36 cultivars. TSUd_3.0 substantively improves prior genome assemblies with new Hi-C and long-read sequence data, covering 531 Mb, containing 41,979 annotated genes and generating a 94.4% BUSCO score. Comparative genomic analysis among select members of the tribe Trifolieae indicated TSUd 3.0 corrects six assembly-error inversion/duplications and confirmed phylogenetic relationships. Its synteny with T. pratense, T. repens, Medicago truncatula and Lotus japonicus genomes were assessed, with the more distantly related T. repens and M. truncatula showing higher levels of co-linearity with Ts than between Ts and its close relative T. pratense. Resequencing of 36 cultivars discovered 7,789,537 SNPs subsequently used for genomic diversity assessment and sequence-based clustering. Heterozygosity estimates ranged from 1% to 21% within the 36 cultivars and may be influenced by admixture. Phylogenetic analysis supported subspecific genetic structure, although it indicates four or five groups, rather than the three recognized subspecies. Furthermore, there were incidences where cultivars characterized as belonging to a particular subspecies clustered with another subspecies when using genomic data. These outcomes suggest that further investigation of Ts sub-specific classification using molecular and morpho-physiological data is needed to clarify these relationships. This upgraded reference genome, complemented with comprehensive sequence diversity analysis of 36 cultivars, provides a platform for future gene functional analysis of key traits, and genome-based breeding strategies for climate adaptation and agronomic performance. Pangenome analysis, more in-depth intra-specific phylogenomic analysis using the Ts core collection, and functional genetic and genomic studies are needed to further augment knowledge of Trifolium genomes.

Genome Sequence and Analysis of Nicotiana benthamiana, the Model Plant for Interactions between Organisms.

Nicotiana benthamiana is widely used as a model plant for dicotyledonous angiosperms. In fact, the strains used in research are highly susceptible to a wide range of viruses. Accordingly, these strains are subject to plant pathology and plant-microbe interactions. In terms of plant-plant interactions, N. benthamiana is one of the plants that exhibit grafting affinity with plants from different families. Thus, N. benthamiana is a good model for plant biology and has been the subject of genome sequencing analyses for many years. However, N. benthamiana has a complex allopolyploid genome, and its previous reference genome is fragmented into 141,000 scaffolds. As a result, molecular genetic analysis is difficult to perform. To improve this effort, de novo whole-genome assembly was performed in N. benthamiana with Hifi reads, and 1,668 contigs were generated with a total length of 3.1 Gb. The 21 longest scaffolds, regarded as pseudomolecules, contained a 2.8-Gb sequence, occupying 95.6% of the assembled genome. A total of 57,583 high-confidence gene sequences were predicted. Based on a comparison of the genome structures between N. benthamiana and N. tabacum, N. benthamiana was found to have more complex chromosomal rearrangements, reflecting the age of interspecific hybridization. To verify the accuracy of the annotations, the cell wall modification genes involved in grafting were analyzed, which revealed not only the previously indeterminate untranslated region, intron and open reading frame sequences but also the genomic locations of their family genes. Owing to improved genome assembly and annotation, N. benthamiana would increasingly be more widely accessible.

Chromosome-scale genome assembly of Eustoma grandiflorum, the first complete genome sequence in the genus Eustoma.

Eustoma grandiflorum (Raf.) Shinn. is an annual herbaceous plant native to the southern United States, Mexico, and the Greater Antilles. It has a large flower with a variety of colors and is an important flower crop. In this study, we established a chromosome-scale de novo assembly of E. grandiflorum genome sequences by integrating four genomic and genetic approaches: (1) Pacific Biosciences (PacBio) Sequel deep sequencing, (2) error correction of the assembly by Illumina short reads, (3) scaffolding by chromatin conformation capture sequencing (Hi-C), and (4) genetic linkage maps derived from an F2 mapping population. Thirty-six pseudomolecules and 64 unplaced scaffolds were created, with a total length of 1,324.8 Mb. A total of 36,619 genes were predicted on the genome as high-confidence genes. A comparison of genome structure between E. grandiflorum and C. canephora or O. pumila suggested whole-genome duplication after the divergence between the families Gentianaceae and Rubiaceae. Phylogenetic analysis with single-copy genes suggested that the divergence time between Gentianaceae and Rubiaceae was 74.94 MYA. Genetic diversity analysis was performed for nine commercial E. grandiflorum varieties bred in Japan, from which 254,205 variants were identified. This first report on the construction of a reference genome sequence in the genus Eustoma is expected to contribute to genetic and genomic studies in this genus and in the family Gentianaceae.

Chromosome-scale genome assembly of a Japanese chili pepper landrace, Capsicum annuum 'Takanotsume'.

Here, we report the genome sequence of a popular Japanese chili pepper landrace, Capsicum annuum 'Takanotsume'. We used long-read sequencing and optical mapping, together with the genetic mapping technique, to obtain the chromosome-scale genome assembly of 'Takanotsume'. The assembly consists of 12 pseudomolecules, which corresponds to the basic chromosome number of C. annuum, and is 3,058.5 Mb in size, spanning 97.0% of the estimated genome size. A total of 34,324 high-confidence genes were predicted in the genome, and 83.4% of the genome assembly was occupied by repetitive sequences. Comparative genomics of linked-read sequencing-derived de novo genome assemblies of two Capsicum chinense lines and whole-genome resequencing analysis of Capsicum species revealed not only nucleotide sequence variations but also genome structure variations (i.e. chromosomal rearrangements and transposon-insertion polymorphisms) between 'Takanotsume' and its relatives. Overall, the genome sequence data generated in this study will accelerate the pan-genomics and breeding of Capsicum, and facilitate the dissection of genetic mechanisms underlying the agronomically important traits of 'Takanotsume'.

Transcriptome analysis of tomato plants following salicylic acid-induced immunity against Clavibacter michiganensis ssp. michiganensis.

Salicylic acid (SA) is known to be involved in the immunity against Clavibacter michiganensis ssp. michiganensis (Cmm) that causes bacterial canker in tomato. To identify the candidate genes associated with SA-inducible Cmm resistance, transcriptome analysis was conducted via RNA sequencing in tomato plants treated with SA. SA treatment upregulated various defense-associated genes, such as PR and GST genes, in tomato cotyledons. A comparison of SA- and Cmm-responsive genes revealed that both SA treatment and Cmm infection commonly upregulated a large number of genes. Gene Ontology (GO) analysis indicated that the GO terms associated with plant immunity were over-represented in both SA- and Cmm-induced genes. The genes commonly downregulated by both SA treatment and Cmm infection were associated with the cell cycle and may be involved in growth and immunity trade-off through cell division. After SA treatment, several proteins that were predicted to play a role in immune signaling, such as resistance gene analogs, Ca2+ sensors, and WRKY transcription factors, were transcriptionally upregulated. The W-box element, which was targeted by WRKYs, was over-represented in the promoter regions of genes upregulated by both SA treatment and Cmm infection, supporting the speculation that WRKYs are important for the SA-mediated immunity against Cmm. Prediction of protein-protein interactions suggested that genes encoding receptor-like kinases and EF-hand proteins play an important role in immune signaling. Thus, various candidate genes involved in SA-inducible Cmm resistance were identified.

Chromosome-scale genome assembly of Glycyrrhiza uralensis revealed metabolic gene cluster centred specialized metabolites biosynthesis.

A high-quality genome assembly is imperative to explore the evolutionary basis of characteristic attributes that define chemotype and provide essential resources for a molecular breeding strategy for enhanced production of medicinal metabolites. Here, using single-molecule high-fidelity (HiFi) sequencing reads, we report chromosome-scale genome assembly for Chinese licorice (Glycyrrhiza uralensis), a widely used herbal and natural medicine. The entire genome assembly was achieved in eight chromosomes, with contig and scaffold N50 as 36.02 and 60.2 Mb, respectively. With only 17 assembly gaps and half of the chromosomes having no or one assembly gap, the presented genome assembly is among the best plant genomes to date. Our results showed an advantage of using highly accurate long-read HiFi sequencing data for assembling a highly heterozygous genome including its complexed repeat content. Additionally, our analysis revealed that G. uralensis experienced a recent whole-genome duplication at approximately 59.02 million years ago post a gamma (γ) whole-genome triplication event, which contributed to its present chemotype features. The metabolic gene cluster analysis identified 355 gene clusters, which included the entire biosynthesis pathway of glycyrrhizin. The genome assembly and its annotations provide an essential resource for licorice improvement through molecular breeding and the discovery of valuable genes for engineering bioactive components and understanding the evolution of specialized metabolites biosynthesis.

De novo whole-genome assembly in an interspecific hybrid table grape, 'Shine Muscat'.

The first genome sequence of an interspecific grape hybrid (Vitis labruscana × Vitis vinifera), 'Shine Muscat', an elite table grape cultivar bred in Japan, is presented. The resultant genome assemblies included two types of sequences: a haplotype-phased sequence of the highly heterozygous genomes and an unphased sequence representing a 'pseudo-haploid' genome. The unphased sequences, assembled to the chromosome level with Hi-C reads, spanned 488.97 Mb in length, 99.1% of the estimated genome size, with 4,595 scaffold sequences and a 23.9-Mb N50 length. The phased sequences had 15,650 scaffolds spanning 1.0 Gb and a 4.2-Mb N50 length. 32,827 high-confidence genes were predicted on the unphased genomes. Clustering analysis of the 'Shine Muscat' gene sequences with three other Vitis species and Arabidopsis indicated that 11,279 orthologous gene clusters were common to Vitis spp. and Arabidopsis, 4,385 were Vitis specific, and 234 were 'Shine Muscat' specific. Whole-genome resequencing was also performed for the parental lines of 'Shine Muscat', Akitsu-21 and 'Hakunan', and parental-specific copy number variations were identified. The obtained genome resources provide new insights that could assist in cultivation and breeding strategies to produce high-quality table grapes.

A genome-wide association and fine-mapping study of white rust resistance in hexaploid chrysanthemum cultivars with a wild diploid reference genome.

White rust caused by Puccinia horiana is one of the most serious diseases of chrysanthemum (Chrysanthemum × morifolium). In this study, we report the DNA markers associated with resistance against P. horiana via a simple approach using the genome of a wild diploid relative, Chrysanthemum seticuspe. First, we identified the important region of the genome in the resistant cultivar "Ariesu" via a genome-wide association study. Simplex single nucleotide polymorphism (SNP) markers mined from ddRAD-Seq were used in a biparental population originating from crosses between resistant "Ariesu" and susceptible "Yellow Queen". The C. seticuspe genome was used as a reference. For the fine mapping of P. horiana resistance locus 2 (Phr2), a comparative whole genome sequencing study was conducted. Although the genome sequences of chrysanthemum cultivars assembled via the short-read approach were fragmented, reliable genome alignments were reconstructed by mapping onto the chromosome level of the C. seticuspe pseudomolecule. Base variants were then identified by comparing the assembled genome sequences of resistant "Ariesu" and susceptible "Yellow Queen". Consequently, SNP markers that were closer to Phr2 compared with ddRAD-Seq markers were obtained. These SNP markers co-segregated with resistance in F1 progenies originating from resistant "Ariesu" and showed robust transferability for detecting Phr2-conferring resistance among chrysanthemum genetic resources. The wild C. seticuspe pseudomolecule, a de facto monoploid genome used for ddRAD-Seq analysis and assembled genome sequence comparison, demonstrated this method's utility as a model for developing DNA markers in hexaploid chrysanthemum cultivars.

Erratum: Comparative QTL mapping for male sterility of cultivated strawberry (Fragaria × ananassa Duch.) using different reference genome sequences.

[This corrects the article DOI: 10.1270/jsbbs.20151.].