Winter rapeseed is not only an important oilseed crop, but also a winter cover crop in Northern China, where its production was severely limited by freezing stress. As an overwinter crop, the production is severely limited by freezing stress. Therefore, understanding the physiological and molecular mechanism of winter rapeseed (Brassica napus L.) in freezing stress responses becomes essential for the improvement and development of freezing-tolerant varieties of Brassica napus. In this study, morphological, physiological, ultrastructure and transcriptome changes in the Brassica napus line “2016TS(G)10” (freezing-tolerance line) that was exposed to –2 °C for 0 h, 1 h, 3 h and 24 h were characterized. The results showed that freezing stress caused seedling dehydration, and chloroplast dilation and degradation. The content of malondialdehyde (MDA), proline, soluble protein and soluble sugars were increased, as well as the relative electrolyte leakage (REL) which was significantly increased at frozen 24 h. Subsequently, RNA-seq analysis revealed a total of 98,672 UniGenes that were annotated in Brassica napus and 3905 UniGenes were identified as differentially expressed genes after being exposed to freezing stress. Among these genes, 2312 (59.21%) were up-regulated and 1593 (40.79%) were down-regulated. Most of these DEGs were significantly annotated in the carbohydrates and energy metabolism, signal transduction, amino acid metabolism and translation. Most of the up-regulated DEGs were especially enriched in plant hormone signal transduction, starch and sucrose metabolism pathways. Transcription factor enrichment analysis showed that the AP2/ERF, WRKY and MYB families were also significantly changed. Furthermore, 20 DEGs were selected to validate the transcriptome profiles via quantitative real-time PCR (qRT-PCR). In conclusion, the results provide an overall view of the dynamic changes in physiology and insights into the molecular regulation mechanisms of winter Brassica napus in response to freezing treatment, expanding our understanding on the complex molecular mechanism in plant response to freezing stress.

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Winter rapeseed (Brassica rapa L.) is the main oilseed crop in northern China and can safely overwinter at 35 (i.e., Tianshui, China) to 48 degrees north latitude (i.e., Altai, Heilongjiang, Raohe, and Xinjiang, China). In order to identify stable reference genes to understand the molecular mechanisms of stress tolerance in winter rapeseed, internal reference genes of winter rapeseed under four abiotic stresses were analyzed using GeNorm, NormFinder, BestKeeper, and RefFinder software. The most stable combinations of internal reference genes were β-actin and SAND in cold-stressed leaves, β-actin and EF1a in cold-stressed roots, F-box and SAND in high temperature-stressed leaves, and PP2A and RPL in high temperature-stressed roots, SAND and PP2A in NaCl-stressed leaves, RPL and UBC in NaCl-stressed roots, RPL and PP2A in PEG-stressed leaves, and PP2A and RPL in PEGstressed roots. Expression profiles of PXG3 were used to verify these results. The stable reference genes identified in this study are useful tools for identifying stress-responsive genes to understand the molecular mechanisms of stress tolerance in winter rapeseed.

Winter rapeseed (Brassica rapa L.) is a major oilseed crop in Northern China, where its production was severely affected by chilling and freezing stress. However, not much is known about the role of differentially accumulated proteins (DAPs) during the chilling and freezing stress. In this study, isobaric tag for relative and absolute quantification (iTRAQ) technology was performed to identify DAPs under freezing stress. To explore the molecular mechanisms of cold stress tolerance at the cellular and protein levels, the morphological and physiological differences in the shoot apical meristem (SAM) of two winter rapeseed varieties, Longyou 7 (cold-tolerant) and Lenox (cold-sensitive), were explored in field-grown plants. Compared to Lenox, Longyou 7 had a lower SAM height and higher collar diameter. The level of malondialdehyde (MDA) and indole-3-acetic acid (IAA) content was also decreased. Simultaneously, the soluble sugars (SS) content, superoxide dismutase (SOD) activity, peroxidase (POD) activity, soluble protein (SP) content, and collar diameter were increased in Longyou 7 as compared to Lenox. A total of 6330 proteins were identified. Among this, 98, 107, 183 and 111 DAPs were expressed in L7 CK/Le CK, L7 d/Le d, Le d/Le CK and L7 d/L7 CK, respectively. Quantitative real-time PCR (RT-qPCR) analysis of the coding genes for seventeen randomly selected DAPs was performed for validation. These DAPs were identified based on gene ontology enrichment analysis, which revealed that glutathione transferase activity, carbohydrate-binding, glutathione binding, metabolic process, and IAA response were closely associated with the cold stress response. In addition, some cold-induced proteins, such as glutathione S-transferase phi 2(GSTF2), might play an essential role during cold acclimation in the SAM of Brassica rapa. The present study provides valuable information on the involvement of DAPs during cold stress responses in Brassica rapa L, and hence could be used for breeding experiments.

Winter and early spring wind soil erosion have considerable impacts on ecosystems, human well-being and agricultural production in the low precipitation zones of northern China. Little is known about the impact of growing winter rapeseed on ecological cropping systems and the associated economic benefits in the wind erosion area. To explore the winter rapeseed cover effect, we conducted a field experiment in which we covered the soil with winter rapeseed, winter wheat and wheat stubble at different plant density levels and used the spring bare ground as the control (CK). The effects of wind erosion, the “winter rapeseed + ” multiple cropping system, and the economic benefits were compared. There was a large difference in the dry matter, the maximum water absorption, the maximum water storage, the soil evaporation and total wind erosion, the amount of sediment transported in the stratum and the wind erosion modulus. Among them, the mean wind erosion modulus of spring sowing bare land was as high as 490.9 kg·hm−2·h−1, which was 7 and 13 times that of winter wheat and winter rapeseed, respectively. As the wind speed increased from 14 to 22 m·s−1, from a small density to a large density, the mean wind erosion modulus decreased from 68 to 17 kg·hm−2·h−1 for winter rapeseed, and 150 to 31 kg·hm−2·h−1 for winter wheat. Total wind-erosion of sediment transport of CK was 18.6 g·m−2 min−1, which was 16 and 31 times the mean value of winter wheat and winter rapeseed, respectively. “Winter rapeseed + ” replanting peanuts, potatoes, rice, seed melons and other crops generally increased the production value by 5–74% compared with wheat and corn intercropping, which was 98–255% higher than the traditional wheat single crop. Our results suggested that the suitable area for planting winter rapeseed in northern China was approximately 3.3 × 106 hm2, and in terms of the best economic and ecological effects, the appropriate density was 5 × 105 plants·hm−2 in northern China. Our results indicated that Chinese winter rapeseed was the best choice for preventing wind erosion and improving ecological and economic benefits in winter and spring in northern China; additionally, winter rapeseed has important impacts on agricultural sustainability in semi-arid and arid climates.