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Summary: High‐quality radish (Raphanus sativus) genome represents a valuable resource for agronomical trait improvements and understanding genome evolution among Brassicaceae species. However, existing radish genome assembly remains fragmentary, which greatly hampered functional genomics research and genome‐assisted breeding. Here, using a NAU‐LB radish inbred line, we generated a reference genome of 476.32 Mb with a scaffold N50 of 56.88 Mb by incorporating Illumina, PacBio and BioNano optical mapping techniques. Utilizing Hi‐C data, 448.12 Mb (94.08%) of the assembled sequences were anchored to nine radish chromosomes with 40 306 protein‐coding genes annotated. In total, 249.14 Mb (52.31%) comprised the repetitive sequences, among which long terminal repeats (LTRs, 30.31%) were the most abundant class. Beyond confirming the whole‐genome triplication (WGT) event in R. sativus lineage, we found several tandem arrayed genes were involved in stress response process, which may account for the distinctive phenotype of high disease resistance in R. sativus. By comparing against the existing Xin‐li‐mei radish genome, a total of 2 108 573 SNPs, 7740 large insertions, 7757 deletions and 84 inversions were identified. Interestingly, a 647‐bp insertion in the promoter of RsVRN1 gene can be directly bound by the DOF transcription repressor RsCDF3, resulting into its low promoter activity and late‐bolting phenotype of NAU‐LB cultivar. Importantly, introgression of this 647‐bp insertion allele, RsVRN1In‐536, into early‐bolting genotype could contribute to delayed bolting time, indicating that it is a potential genetic resource for radish late‐bolting breeding. Together, this genome resource provides valuable information to facilitate comparative genomic analysis and accelerate genome‐guided breeding and improvement in radish. [ABSTRACT FROM AUTHOR]

CLAVATA3/EMBRYO SURROUNDING REGION-related (CLE) peptides are a class of small molecules involved in plant growth and development. Although radish (Raphanus sativus) is an important root vegetable crop worldwide, the functions of CLE peptides in its taproot formation remain elusive. Here, a total of 48 RsCLE genes were identified from the radish genome. RNA in situ hybridization showed that RsCLE22a gene was highly expressed in the vascular cambium. Overexpression of RsCLE22a inhibited root growth by impairing stem cell proliferation in Arabidopsis, and radish plants with exogenous supplementation of RsCLE22 peptide (CLE22p) showed a similar phenotype. The vascular cambial activity was increased in RsCLE22a -silenced plants. Transcriptome analysis revealed that CLE22p altered the expression of several genes involved in meristem development and hormone signal transduction in radish. Immunolocalization results showed that CLE22p increased auxin accumulation in vascular cambium. Yeast one-hybrid and dual-luciferase assays showed that the WUSCHEL-RELATED HOMEOBOX 4 (RsWOX4) binds to RsCLE22a promoter and activates its transcription. The expression level of RsWOX4 was related to vascular cambial activity and was regulated by auxin. Furthermore, a RsCLE22a–RsWOX4 module is proposed to regulate taproot vascular cambium activity through an auxin signaling-related pathway in radish. These findings provide novel insights into the regulation of root growth in a horticultural crop. [ABSTRACT FROM AUTHOR]

Radish (Raphanus sativus L.), an important annual or biennial root vegetable crop, is widely cultivated in the world for its high nutritive value. Isolated microspore culture (IMC) is one of the most effective methods for rapid development of homozygous lines. Due to imperfection of the IMC technology system, it is particularly important to establish an efficient IMC system in radish. In this study, the effects of different factors on radish microspore embryogenesis were investigated with 23 genotypes. Buds with the largest population of late-uninucleate-stage microspores were most suitable for embryogenesis, with a ratio of petal length to anther length (P/A) in buds of about 3/4 ~ 1. Cold pretreatment was found to be genotype specific, and the highest microspore-derived embryoid (MDE) yield occurred for treatment of the heat shock of 48 h. In addition, the supplement of 0.75 g/L activated charcoal (AC) could increase the yield of embryoids. It was found that genotypes, bud size, as well as temperature treatments had significant effects on microspore embryogenesis. Furthermore, somatic embryogenesis–related kinase (SERK) genes were profiled by reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis, which indicated that they are involved in the process of MDE formation and plantlet regeneration. The ploidy of microspore-derived plants was identified by chromosome counting and flow cytometry, and the microspore-derived plants were further proved as homozygous plants through expressed sequence tags-simple sequence repeats (EST-SSR) and genetic-SSR markers. The results would facilitate generating the large-scale double haploid (DH) from various genotypes, and promoting further highly efficient genetic improvement in radish. [ABSTRACT FROM AUTHOR]

• Totally 42 members of the Histone gene family were firstly identified in radish. • Seven RsCENH3 genes exhibited specifically high expression level in the radish bud. • RsCENH3 gene might be involved in promoting microspore development in radish. Histone, a predominant protein component of chromatin, participates in DNA packaging and transcriptional regulation. However, the available information of Histone gene family is limited in radish. In this study, a total of 42 Histone gene family members were identified from the radish genome. Sequence alignment and phylogenetic analyses classified the Histone family into three groups (H2A, H2B and H3). Motif analysis showed that the functions of some motifs shared by H3 subfamily genes were related to chromosome regulation and cell development activities, such as motif 5 containing Cks1 and PPR region. Analysis of intron/exon structure indicated that RsCENH3 (RsHistone 18) has the characteristics of variant Histone. Furthermore, several motifs, including the LTR, G-box and TC-elements, were found in the promoters of RsHistone genes, which involved in cell development or various abiotic stresses responses. Transcriptome analysis indicated that the RsHistone genes exhibited higher expression level in floral buds than in roots and leaves. Subcellular localization showed that the RsCENH3 was localized on the nucleus, and it was highly expressed in the floral bud of 3.0–4.0 mm in radish. These findings would provide valuable information for characterization and potential utilization of Histone genes, and facilitate the efficient induction of double haploid plants in radish. [ABSTRACT FROM AUTHOR]

• The identified 65 Aux/IAA genes show an uneven chromosomal distribution in radish. • Six Aux/IAA genes exhibit specifically high expression level in radish root tissues. • RsIAA33 gene might involve in promoting taproot thickening and development in radish. Auxin/indole–3–acetic acid (Aux/IAA) genes encode short lived nuclear proteins that cooperated with auxin or auxin response factor (ARF), which are involved in plant growth and developmental processes. However, it's still ambiguous how the Aux/IAA genes regulate the process governing taproot thickening in radish. Herein, 65 Aux/IAA genes were identified from the radish genome. Gene duplication analysis showed that two pairs of tandem duplication and 17 (27%) segmental duplication events were identified among Aux/IAA family genes in radish. Transcriptomic analysis revealed that most of Aux/IAA genes (52/65) exhibited differential expression pattern in different root tissues, and six root–specific genes were highly expressed in root cortex, cambium, xylem, and root tip in radish. RT–qPCR analysis showed that the expression level of RsIAA33 was the highest at cortex splitting stage (CSS), and early expanding stage (ES). Furthermore, amiRNA–mediated gene silencing of RsIAA33 indicated that it could inhibit the reproductive growth, thus promoting taproot thickening and development. These results would provide valuable information for elucidating the molecular function of Aux/IAA genes involved in taproot thickening in radish. [ABSTRACT FROM AUTHOR]