Background: The basic helix–loop–helix (bHLH) is the second largest gene family in the plant, some members play important roles in pistil development and response to drought, waterlogging, cold stress and salt stress. The bHLH gene family has been identified in many species, except forBrassica oleraceaandB. napusthus far.This study aims to identify the bHLH family members inB. oleracea,B. rapaandB. napus, and elucidate the expression, duplication, phylogeny and evolution characters of them.Result: A total of 268 bHLH genes inB. oleracea, 440 genes inB. napus, and 251 genes inB. rapa, including 21 new bHLH members, have been identified.Subsequently, the analyses of the phylogenetic trees, conserved motifs and gene structures showed that the members in the same subfamily were highly conserved. MostKa/Ksvalues of homologous gene were < 1, which indicated that these genes suffered from strong purifying selection for retention. The retention rates ofBrabHLHandBolbHLHgenes were 51.6 and 55.1%, respectively. The comparative expression patterns betweenB. rapaandB. napusshowed that they had similar expression patterns in the root and contrasting patterns in the stems, leaves, and reproductive tissues. In addition, there were 41 and 30 differential expression bHLH genes under the treatments of ABA and JA, respectively, and the number of down regulation genes was significantly more than up regulation genes.Conclusion: In the present study, we identified and performed the comparative genomics analysis of bHLH gene family amongB. oleracea,B. rapaandB. napus, and also investigated their diversity. The expression patterns betweenB. rapaandB. napusshows that they have the similar expression pattern in the root and opposite patterns in the stems, leaves, and reproduction tissues. Further analysis demonstrated that some bHLH gene members may play crucial roles under the abiotic and biotic stress conditions. This is the first to report on the bHLH gene family analysis inB. oleraceaandB. napus,which can offer useful information on the functional analysis of the bHLH gene in plants.

Plasmodiophora brassicae (P. brassicae) is a soil-born pathogen worldwide and can infect most cruciferous plants, which causes great yield decline and economic losses. It is not well known how microbial diversity and community composition change during P. brassicae infecting plant roots. Here, we employed a resistant and a susceptible pakchoi cultivar with and without inoculation with P. brassicae to analyze bacterial and fungal diversity using 16S rRNA V3-V4 and ITS_V1 regions, respectively. 16S rRNA V3-V4 and ITS_V1 regions were amplified and sequenced separately. Results revealed that both fungal and bacterial diversity increased, and composition was changed in the rhizosphere soil of the susceptible pakchoi compared with the resistant cultivar. In the four groups of R_mock, S_mock, R_10d, and S_10d, the most relatively abundant bacterium and fungus was Proteobacteria, accounting for 61.92%, 58.17%, 48.64%, and 50.00%, respectively, and Ascomycota, accounting for 75.11%, 63.69%, 72.10%, and 90.31%, respectively. A total of 9488 and 11,914 bacteria were observed uniquely in the rhizosphere soil of resistant and susceptible pakchoi, respectively, while only 80 and 103 fungi were observed uniquely in the correlated soil. LefSe analysis showed that 107 and 49 differentially abundant taxa were observed in bacteria and fungi. Overall, we concluded that different pakchoi cultivars affect microbial diversity and community composition, and microorganisms prefer to gather around the rhizosphere of susceptible pakchoi. These findings provide a new insight into plant–microorganism interactions.

Downy mildew caused by the obligate parasite Hyaloperonospora brassicae is a devastating disease for Brassica species. Infection of Hyaloperonospora brassicae often leads to yellow spots on leaves, which significantly impacts quality and yield of pakchoi. In the present study, we conducted a comparative transcriptome between the resistant and susceptible pakchoi cultivars in response to Hyaloperonospora brassicae infection. A total of 1073 disease-resistance-related differentially expressed genes were identified using a Venn diagram. The Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that these genes were mainly involved in plant−pathogen interaction, plant hormone signal transduction, and other photosynthesis-related metabolic processes. Analysis of the phytohormone content revealed that salicylic acid increased significantly in the resistant material after inoculation with Hyaloperonospora brassicae, whereas the contents of jasmonic acid, abscisic acid, and 1-aminocyclopropane-1-carboxylic acid decreased. Exogenous salicylic acid treatment also significantly upregulated Hyaloperonospora brassicae-induced genes, which further confirmed a crucial role of salicylic acid during pakchoi defense against Hyaloperonospora brassicae. Based on these findings, we suggest that the salicylic-acid-mediated signal transduction contributes to the resistance of pakchoi to downy mildew, and PAL1, ICS1, NPR1, PR1, PR5, WRKY70, WRKY33, CML43, CNGC9, and CDPK15 were involved in this responsive process. Our findings evidently contribute to revealing the molecular mechanism of pakchoi defense against Hyaloperonospora brassicae.

Simple sequence repeats (SSRs), also known as microsatellites, stand out as the most crucial molecular markers in both animals and plants owing to their high polymorphism, extensive information content, ease of detection through polymerase chain reaction (PCR) assays, and widespread distribution across the genome. In this study, a total of 125,443 SSR loci were identified from the whole-genome sequence of B. oleracea, and 82,948 primer pairs for SSR have been designed. Furthermore, each primer pair is designated with a unique identifier (ranging from BolSSR00001 to BolSSR82984). Our findings indicated that certain markers within them could be transferred to other cruciferous crops. In addition, a total of 336 pairs of SSR primers have been used to screen the polymorphism between the bolting-resistant and bolting-easy gene pools. After the test of verification with F₂ generation individual plants, we obtained an SSR dominant marker, BolSSR040196, linked with bolting-resistant locus in cabbage, and the genetic distance between this SSR marker and the bolting-resistant locus was 10.69 cM. Moreover, BolSSR040196 is located on the C05 chromosome with a CT motif, characterized by a repeat of 9 in bolting-easy plants and 11 in bolting-resistant plants. Haplotype analysis showed that the correct prediction rate reached 82.35%. The BolSSR040196 marker can be used in marker-assisted selection (MAS) breeding, offering a straightforward and efficient approach for bolting-resistant cabbage breeding in the future.

Hypocotyl length is a critical determinant for the efficiency of mechanical harvesting in pakchoi production, but the knowledge on the molecular regulation of hypocotyl growth is very limited. Here, we report a spontaneous mutant of pakchoi, lhy7.1, and identified its characteristics. We found that it has an elongated hypocotyl phenotype compared to the wild type caused by the longitudinal growth of hypocotyl cells. Different light quality treatments, transcriptome, and proteomic analyses were performed to reveal the molecular mechanisms of hypocotyl elongation. The data showed that the hypocotyl length of lhy7.1 was significantly longer than that of WT under red, blue, and white lights but there was no significant difference under dark conditions. Furthermore, we used transcriptome and label-free proteome analyses to investigate differences in gene and protein expression levels between lhy7.1 and WT. At the transcript level, 4568 differentially expressed genes (DEGs) were identified, which were mainly enriched in “plant hormone signal transduction”, “photosynthesis”, “photosynthesis–antenna proteins”, and “carbon fixation in photosynthetic organisms” pathways. At the protein level, 1007 differentially expressed proteins (DEPs) were identified and were mainly enriched in photosynthesis-related pathways. The comprehensive transcriptome and proteome analyses revealed a regulatory network of hypocotyl elongation involving plant hormone signal transduction and photosynthesis-related pathways. The findings of this study help elucidate the regulatory mechanisms of hypocotyl elongation in lhy7.1.

Clubroot disease is a kind of soil-borne disease that seriously infects Brassica species. In this study, we collected 121 varieties of non-heading Chinese cabbages. In order to better understand the genetic variation and to screen suitable clubroot disease-resistant parental material, we re-sequenced them to examine the population genetic structure, population genetic diversity, population differentiation index, and selective sweep based on SNPs. The mapping rate with the reference genome was high, and data quality analysis revealed that the sequencing quality was good. The annotated data indicated that intronic and intergenic areas held the majority of SNPs and indels. Four subgroups of 121 non-heading Chinese cabbages were identified using principal component analysis, phylogenetic tree, and genetic structure analysis. An examination of genetic diversity revealed that while selfing may happen in subgroups C and D, heterozygosity may exist in subgroups A and B. In subgroup B, self-fertilization is not possible. There was a moderate degree of genetic differentiation between subgroups B and C (Fst = 0.0744347). For genes in certain sweep regions, we also ran GO enrichment and KEGG enrichment analysis. Two disease resistance-related genes, BraA01g042910.3.5C and BraA06g019360.3.5C, were examined. These findings will serve as a theoretical foundation for developing novel, clubroot disease-resistant types of non-heading Chinese cabbages.