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Brassica juncea var. tumida Tsen et Lee (Tumorous stem mustard) is an unique vegetable in China. Its enlarged tumorous stem was used as main raw material to produce pickle (Zhacai). In practice, early-bolting happens around 15% of planting area all year and inhibits its production. Here, about 209 PP2C proteins were identified through HMMER software and divided into 13 sub-families in B. juncea. BjuPP2C52 belongs to E sub-family, was up-regulated at reproductive growth stages and interacts with BjuFKF1, a key protein in regulating plant photoperiod flowering, in vitro and in vivo. To explore interactive proteins, BjuPP2C52 was used as bait, 12 potential interactive proteins were screened from yeast library, and they are BjuCOL3, BjuCOL5, BjuAP2, BjuAP2-1, BjuSVP-1, BjuFLC-2, BjuSKP1f, BjuA014572, BjuA008686, BjuO002119, BjuB036787 and BjuA019268. Further study verified that 10 out of the 12 screened proteins interacted with BjuPP2C52 in vivo. qRT-PCR was conducted to understand the expression pattern of those 10 interactive proteins in different tissues and development stages in B. juncea. The results showed that BjuCOL3, BjuCOL5, BjuB036787 and BjuA019268 were significantly up-regulated, while BjuA008686 and BjuO002119 were down-regulated in flowers compared with other four tissues. In developmental stages, BjuCOL5, BjuAP2, BjuAP2-1, BjuA014572, BjuB036787 and BjuA019268 were significantly up-regulated, while BjuSVP-1, BjuA008686 and BjuO002119 were down-regulated at reproductive stages. Based on the results, BjuCOL5, BjuAP2, BjuAP2-1, BjuSVP-1, BjuA014572, BjuB036787 and BjuA019268 may function in regulating flowering time in B. juncea.

Radish (Raphanus sativus L.), a biennial root vegetable crop belonging to the Brassicaceae family, is an economically important vegetable grown worldwide. Radish is a recalcitrant variety for embryogenic callus induction that does not result in an efficient genetic transformation system. In this study, an efficiency regeneration system of callus induction and bud differentiation in radish was established using cotyledons explants. Among the 13 types of explants (epicotyl, cotyledon leaves, cotyledons with petioles, petioles of cotyledons, euphylls, petioles of euphylls, ovaries, pericarps, immature embryos, mature embryos, stems, stems with buds, and terminal buds), cotyledon leaves had the highest induction rate (91.01%) and were the most suitable explant. In addition, the optimal conditions for callus induction was MS-1 medium supplemented with 0.2 mg/L 2,4-dichlorophenoxyacetic acid, 0.5 mg/L thidiazuron (TDZ), 0.2 mg/L α-naphthalene acetic acid (NAA), 4 mg/L 6-benzylaminopurine (6-BA), 0.1 mg/L kinetin, 8 g/L agar, and 3 g/L sugar, with a callus induction rate of 94.77%. Moreover, the best medium for shoot regeneration was MS-1 medium supplemented with 1 mg/L TDZ, 1 mg/L 6-BA, 8 g/L agar, and 3 g/L sugar, with a bud regeneration rate of 15.54%. The most suitable medium of rooting was 1/2 MS-1 medium fortified with 0.2 mg/L NAA, 0.2 mg/L 3-indolebutyric acid, 0.1% activated carbon, 8 g/L agar, and 3 g/L sugar, achieving a rooting ratio of 89.05% and transplanting survival rate survival ratio of 94.92%. This study lays a good foundation for future breeding and molecular studies of radishes.

Tumorous stem mustard is well known for its swollen stem from which pickled “Fuling Mustard” is made. The molecular mechanisms governing the formation of the modified swollen stems are still poorly understood. This paper aims to identify candidate genes involved in the developmental regulation of the swollen stems. We sought to map previously published transcriptome datasets for Brassica juncea, including those derived from swollen stems at four different developmental stages and a mutant variety without swollen stems. Using pairwise comparisons of the five datasets, we identified 31 368 differentially expressed genes (DEGs). A total of 55 DEGs related to plant hormone signal transduction and 259 continuously up- or downregulated transcription factors were identified during stem development using Gene Ontology (GO) analysis. Quantitative real-time PCR (qRT-PCR) results showed that the expressions of 12 important candidate DEGs were consistent with RNA-seq results. Our study provided digital gene expression profiling and a dynamic view of the swollen stem development process. Furthermore, we identified candidate genes for further studies on mechanisms of modified stem development in non-model species.

Abscisic acid (ABA) plays crucial roles in plant response to environmental stresses and development. The clade A phosphatases (PP2Cs) play a crucial role in ABA signaling. However, little is known about the details regarding PP2Cs family genes in Brassica juncea var. tumida. Here, 20 clade A PP2Cs family genes were identified in tuber mustard genome, including BjuABI1s, BjuABI2s, BjuAHG1s, BjuAHG3s, BjuHAB1, BjuHAB2s, BjuHAI1s, BjuHAI2s and BjuHAI3. The promoters of BjuPP2Cs family genes contained various of cis-acting elements, such as ABRE, GT1GMSCAM4, ARFAT and MYB1AT. We also analyzed the expression pattern of clade A BjuPP2Cs under abiotic stresses (low temperature, NaCl and ABA) treatment, pathogen Plasmodiophora brassicae treatment and different stages of stem swollen. The results suggested that clade A BjuPP2Cs regulated tuber mustard response to P. brassicae to mediate the formation of clubroot and might play roles in stem swelling and response to abiotic stresses. This study provides valuable information for further functional investigations of clade A PP2Cs family genes in B. juncea var. tumida.

The swollen succulent stem of tumorous stem mustard (Brassica juncea var. tumida Tsen et Lee) is the raw material in the processing of Chinese Fuling zhàcài. We found that total polyphenols and flavonoids contents of tumorous stem mustard leaf extracts (TSMLE) were significant higher than that of tumorous stem mustard stem extracts (TSMSE). The percentage of isorhamnetin 5-O-hexoside, methyl quercetin O-hexoside, and luteolin O-hexosyl-O-hexosyl-O-hexoside in TSMLE were 170, 230 and 694 times higher than that in TSMSE. TSMLE presented stronger antioxidant capacity against 2,2'-Azino-bis[3-ethylbenzothiazoline]-6-sulfonic acid cationic free radical (ABTS+) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radicals, and it also showed higher reducing power and ferric reducing antioxidant power. Both TSMLE and TSMSE inhibited lung carcinoma cell growth in a dose-dependent manner, while TSMLE was more effective against A549 cells than TSMSE. This is the first report indicates that tumorous stem mustard, especially its leaf, is a potential diet source in preventing of oxidative stress and cancer.

The Salt Overly Sensitive (SOS) pathway in Arabidopsis thaliana plays important roles in maintaining appropriate ion homeostasis in the cytoplasm and regulating plant tolerance to salinity. However, little is known about the details regarding SOS family genes in the tuber mustard crop (Brassica juncea var. tumida). Here, 12 BjSOS family genes were identified in the B. juncea var. tumida genome including two homologous genes of SOS1, one and three homologs of SOS2 and SOS3, two homologs of SOS4, two homologs of SOS5 and two homologs of SOS6, respectively. The results of conserved motif analysis showed that these SOS homologs contained similar protein structures. By analyzing the cis-elements in the promoters of those BjSOS genes, several hormone- and stress-related cis-elements were found. The results of gene expression analysis showed that the homologous genes were induced by abiotic stress and pathogen. These findings indicate that BjSOS genes play crucial roles in the plant response to biotic and abiotic stresses. This study provides valuable information for further investigations of BjSOS genes in tuber mustard.

Abscisic acid (ABA) plays important roles in multiple physiological processes, such as plant response to stresses and plant development. The ABA receptors pyrabactin resistance (PYR)/ PYR1-like (PYL)/regulatory components of ABA receptor (RCAR) play a crucial role in ABA perception and signaling. However, little is known about the details regarding PYL family genes in Brassica juncea var. tumida. Here, 25 PYL family genes were identified in B. juncea var. tumida genome, including BjuPYL3, BjuPYL4s, BjuPYL5s, BjuPYL6s, BjuPYL7s, BjuPYL8s, BjuPYL10s, BjuPYL11s, and BjuPYL13. The results of phylogenic analysis and gene structure showed that the PYL family genes performed similar gene characteristics. By analyzing cis-elements in the promoters of those BjuPYLs, several hormone and stress related cis-elements were found. The results of gene expression analysis showed that the ABA receptor homologous genes were induced by abiotic and biotic stress. The tissue-specific gene expression patterns of BjuPYLs also suggested those genes might regulate the stem swelling during plant growth. These findings indicate that BjuPYLs are involved in plant response to stresses and organ development. This study provides valuable information for further functional investigations of PYL family genes in B. juncea var. tumida.

Stem mustard (Brassica juncea var. tumida Tsen et Lee) is one of a unique vegetable in China and the tumor stem is its economic product. With the expansion planting area to high altitude in Fuling, early-bolting has become the most critical issue in its cultivation. Previous studies found that stem mustard early-bolting in high latitude closely related to short waves. FKF1 (FLAVIN BINDING, KELCH REPEAT, F-BOX1) is a plant-specific LOV blue light receptor, mainly regulates photoperiodic flowering and cycle rhythm. We used BjuFKF1 as the bait protein and detected its interaction protein BjuPP2C52. Further, GST pull down assay confirmed the interaction in vitro. Yeast two hybrid assay showed that both the F-box domain and KELCH repeat domain of BjuFKF1 interacted with BjuPP2C52 in vivo. RT-PCR and qRT-PCR was used to analyze gene expression in different tissues and development stages, respectively. RT-PCR results indicated that BjuFKF1 was expressed in tumorous stem at an extremely high level, while BjuPP2C52 was highly expressed in both flowers and tumorous stem. qRT-PCR assays showed that both BjuFKF1 and BjuPP2C52 were up-regulated expressed in the reproductive growth stages. Sub-cellular assays showed that BjuFKF1 and BjuPP2C52 located in the nucleus and plasma membrane, respectively. Taken together, our results demonstrated that BjuFKF1 interacted with BjuPP2C52 in vitro and vivo, BjuFKF1 was expressed at an extremely high level in tumorous stem, BjuPP2C52 was highly expressed in flowers. BjuFKF1 and BjuPP2C52 were up-regulated expressed in reproductive growth stages. Sub-cellular assays showed that BjuFKF1 and BjuPP2C52 located in the nucleus and cell membrane, respectively. It was speculated that BjuFKF1 may through the pathway of “BjuFKF1-BjuPP2C52-BjuAGB1-BjuNH3-Bjuphot1” to regulate stem mustard response to blue light and eventually bolting and flowering time.