Rapeseed (Brassica napus) is one of the important oil crops worldwide. Its production is often threatened by drought stress. Here, we identify a transcription factor (BnaA9.NF-YA7) that negatively regulates drought tolerance through genome-wide association study in B. napus. The presence of two SNPs within a CCAAT cis element leads to downregulation of BnaA9.NF-YA7 expression. In addition, the M63I (G-to-C) substitution in the transactivation domain can activate low level expression of BnaA4.DOR, which is an inhibitory factor of ABA-induced stomatal closure. Furthermore, we determine that Bna.ABF3/4s directly regulate the expression of BnaA9.NF-YA7, and BnaA9.NF-YA7 indirectly suppresses the expression of Bna.ABF3/4s by regulation of Bna.ASHH4s. Our findings uncover that BnaA9.NF-YA7 serves as a supplementary role for ABA signal balance under drought stress conditions, and provide a potential molecular target to breed drought-tolerant B. napus cultivars.

Background: Soybean is a globally important legume crop that provides a primary source of high-quality vegetable protein and oil. Seed protein content (SPC) is a valuable quality trait controlled by multiple genes in soybean. Results: In this study, we performed quantitative trait loci (QTL) mapping, QTL-seq, and RNA sequencing (RNA-seq) to reveal the genes controlling protein content in the soybean by using the high protein content variety Nanxiadou 25. A total of 50 QTL for SPC distributed on 14 chromosomes except chromosomes 4, 12, 14, 17, 18, and 19 were identified by QTL mapping using 178 recombinant inbred lines (RILs). Among these QTL, the major QTL qSPC_20–1 and qSPC_20–2 on chromosome 20 were repeatedly detected across six tested environments, corresponding to the location of the major QTL detected using whole-genome sequencing-based QTL-seq. 329 candidate DEGs were obtained within the QTL region of qSPC_20–1 and qSPC_20–2 via gene expression profile analysis. Nine of which were associated with SPC, potentially representing candidate genes. Clone sequencing results showed that different single nucleotide polymorphisms (SNPs) and indels between high and low protein genotypes in Glyma.20G088000 and Glyma.16G066600 may be the cause of changes in this trait. Conclusions: These results provide the basis for research on candidate genes and marker-assisted selection (MAS) in soybean breeding for seed protein content.

APETALA2-like genes encode plant-specific transcription factors, some of which possess one microRNA172 (miR172) binding site. The miR172 and its target euAP2 genes are involved in the process of phase transformation and flower organ development in many plants. However, the roles of miR172 and its target AP2 genes remain largely unknown in Brassica napus (B. napus). In this study, 19 euAP2 and four miR172 genes were identified in the B. napus genome. A sequence analysis suggested that 17 euAP2 genes were targeted by Bna-miR172 in the 3′ coding region. EuAP2s were classified into five major groups in B.napus. This classification was consistent with the exon-intron structure and motif organization. An analysis of the nonsynonymous and synonymous substitution rates revealed that the euAP2 genes had gone through purifying selection. Whole genome duplication (WGD) or segmental duplication events played a major role in the expansion of the euAP2 gene family. A cis-regulatory element (CRE) analysis suggested that the euAP2s were involved in the response to light, hormones, stress, and developmental processes including circadian control, endosperm and meristem expression. Expression analysis of the miR172-targeted euAP2s in nine different tissues showed diverse spatiotemporal expression patterns. Most euAP2 genes were highly expressed in the floral organs, suggesting their specific functions in flower development. BnaAP2–1, BnaAP2–5 and BnaTOE1–2 had higher expression levels in late-flowering material than early-flowering material based on RNA-seq and qRT-PCR, indicating that they may act as floral suppressors. Overall, analyses of the evolution, structure, tissue specificity and expression of the euAP2 genes were peformed in B.napus. Based on the RNA-seq and experimental data, euAP2 may be involved in flower development. Three euAP2 genes (BnaAP2–1, BnaAP2–5 and BnaTOE1–2) might be regarded as floral suppressors. The results of this study provide insights for further functional characterization of the miR172 /euAP2 module in B.napus.

Background: Brassica napus is one of the most important oilseed crops, and also an important biofuel plant due to its low air pollution and renewability. Growth period are important traits that affect yield and are crucial for its adaptation to different environments in B. napus. Results: To elucidate the genetic basis of growth period traits, genome-wide association analysis (GWAS) and linkage mapping were employed to detect the quantitative trait loci (QTL) for days to initial flowering (DIF), days to final flowering (DFF), flowering period (FP), maturity time (MT), and whole growth period (GP). A total of 146 SNPs were identified by association mapping, and 83 QTLs were identified by linkage mapping using the RIL population. Among these QTLs, 19 were pleiotropic SNPs related to multiple traits, and six (q18DFF.A03-2, q18MT.A03-2, q17DFF.A05-1, q18FP.C04, q17DIF.C05 and q17GP.C09) were consistently detected using both mapping methods. Additionally, we performed RNA sequencing to analyze the differential expression of gene (DEG) transcripts between early- and late-flowering lines selected from the RIL population, and the DEGs were integrated with association mapping and linkage analysis to confirm their roles in the growth period. Consequently, 12 candidate genes associated with growth period traits were identified in B. napus. Among these genes, seven have polymorphic sites in the coding sequence and the upstream 2-kb sequence based on the resequencing data. The haplotype BnaSOC1.A05-Haplb and BnaLNK2.C06-Hapla showed more favorable phenotypic traits. Conclusions: The candidate genes identified in this study will contribute to our genetic understanding of growth period traits and can be used as targets for target mutations or marker-assisted breeding for rapeseed adapted to different environments.

Soil is indispensable for agricultural production but has been seriously polluted by cadmium and salt in recent years. Many crops are suffering from this, including rapeseed, the third largest global oilseed crop. However, genes simultaneously related to both cadmium and salt stress have not been extensively reported yet. In this study, BnaA10.WRKY75 was screened from previous RNA-seq data related to cadmium and salt stress and further analyses including sequence comparison, GUS staining, transformation and qRT-PCR were conducted to confirm its function. GUS staining and qRT-PCR results indicated BnaA10.WRKY75 was induced by CdCl₂ and NaCl treatment. Sequence analysis suggested BnaA10.WRKY75 belongs to Group IIc of the WRKY gene family and transient expression assay showed it was a nuclear localized transcription factor. BnaA10.WRKY75-overexpressing Arabidopsis and rapeseed plants accumulated more H₂O₂ and O₂⁻ and were more sensitive to CdCl₂ and NaCl treatment compared with untransformed plants, which may be caused by the downregulation of BnaC03.CAT2. Our study reported that BnaA10.WRKY75 increases sensitivity to cadmium and salt stress by disrupting the balance of reactive oxygen species both in Arabidopsis and rapeseed. The results support the further understanding of the mechanisms underlying cadmium and salt tolerance and provide BnaA10.WRKY75 as a valuable gene for rapeseed abiotic stress breeding.