Physiological and Transcriptome Analysis on Diploid and Polyploid Populus ussuriensis Kom. under Salt Stress.

Populus ussuriensis Kom. is a valuable forest regeneration tree species in the eastern mountainous region of Northeast China. It is known that diploid P. ussuriensis (CK) performed barely satisfactorily under salt stress, but the salt stress tolerance of polyploid (i.e., triploid (T12) and tetraploid (F20)) P. ussuriensis is still unknown. In order to compare the salt stress tolerance and salt stress response mechanism between diploid and polyploid P. ussuriensis, phenotypic observation, biological and biochemistry index detections, and transcriptome sequencing (RNA-seq) were performed on CK, T12, and F20. Phenotypic observation and leaf salt injury index analysis indicated CK suffered more severe salt injury than T12 and F20. SOD and POD activity detections indicated the salt stress response capacity of T12 was stronger than that of CK and F20. MDA content, proline content and relative electric conductivity detections indicated CK suffered the most severe cell-membrane damage, and T12 exhibited the strongest osmoprotective capacity under salt stress. Transcriptome analysis indicated the DEGs of CK, T12, and F20 under salt stress were different in category and change trend, and there were abundant WRKY, NAM, MYB and AP2/ERF genes among the DEGs in CK, T12, and F20 under salt stress. GO term enrichment indicated the basic growth progresses of CK, and F20 was obviously influenced, while T12 immediately launched more salt stress response processes in 36 h after salt stress. KEGG enrichment indicated the DEGs of CK mainly involved in plant−pathogen interaction, ribosome biogenesis in eukaryotes, protein processing in endoplasmic reticulum, degradation of aromatic compounds, plant hormone signal transduction, photosynthesis, and carbon metabolism pathways. The DEGs of T12 were mainly involved in plant−pathogen interaction, cysteine and methionine metabolism, phagosomes, biosynthesis of amino acids, phenylalanine, tyrosine and tryptophan biosynthesis, plant hormone signal transduction, and starch and sucrose metabolism pathways. The DEGs of F20 were mainly involved in plant hormone signal transduction, plant−pathogen interaction, zeatin biosynthesis, and glutathione metabolism pathways. In conclusion, triploid exhibited stronger salt stress tolerance than tetraploid and diploid P. ussuriensis (i.e., T12 > F20 > CK). The differences between the DEGs of CK, T12, and F20 probably are the key clues for discovering the salt stress response signal transduction network in P. Ussuriensis.

Comparative Transcriptomics and Proteomics Analyses of Leaves Reveals a Freezing Stress-Responsive Molecular Network in Winter Rapeseed (Brassica rapa L.).

Winter rapeseed is susceptible to low temperature during winter in Northwest China, which could lead to a severe reduction of crop production. The freezing temperature could stress the whole plant, especially the leaf, and ultimately harm the survival rate of winter rapeseed. However, the molecular mechanism underlying freezing tolerance is still unclear in winter rapeseed. In this study, a comprehensive investigation of winter rapeseed freezing tolerance was conducted at the levels of transcript, protein, and physiology and biochemistry, using a pair of freezing-sensitive and freezing-resistant cultivars NQF24 and 17NTS57. There were 4,319 unique differentially expressed genes (DEGs) and 137 unique differentially abundant proteins (DAPs) between two cultivars identified in leaf under freezing stress. Function enrichment analysis showed that most of the enriched DEGs and DAPs were involved in plant hormone signal transduction, alpha-linolenic/linoleic acid metabolism, peroxisome, glutathione metabolism, fatty acid degradation, and secondary metabolite biosynthesis pathways. Based on our findings, it was speculated that freezing tolerance formation is caused by increased signal transduction, enhanced biosynthesis of protein, secondary metabolites, and plant hormones, elevated energy supply, greater reactive oxygen species scavenging, and lower lipid peroxidation as well as stronger cell stability in leaf under freezing stress. These results provide a comprehensive profile of leaf response under freezing stress, which have potential to be used as selection indicators of breeding programs to improve freezing tolerance in rapeseed.

Identification of differentially expressed genes involved in amino acid and lipid accumulation of winter turnip rape (Brassica rapa L.) in response to cold stress.

Winter turnip rape (Brassica rapa L.) is an important overwintering oil crop that is widely planted in northwestern China. It considered to be a good genetic resource for cold-tolerant research because its roots can survive harsh winter conditions. Here, we performed comparative transcriptomics analysis of the roots of two winter turnip rape varieties, Longyou7 (L7, strong cold tolerance) and Tianyou2 (T2, low cold tolerance), under normal condition (CK) and cold stress (CT) condition. A total of 8,366 differentially expressed genes (DEGs) were detected between the two L7 root groups (L7CK_VS_L7CT), and 8,106 DEGs were detected for T2CK_VS_T2CT. Among the DEGs, two ω-3 fatty acid desaturase (FAD3), two delta-9 acyl-lipid desaturase 2 (ADS2), one diacylglycerol kinase (DGK), and one 3-ketoacyl-CoA synthase 2 (KCS2) were differentially expressed in the two varieties and identified to be related to fatty acid synthesis. Four glutamine synthetase cytosolic isozymes (GLN), serine acetyltransferase 1 (SAT1), and serine acetyltransferase 3 (SAT3) were down-regulated under cold stress, while S-adenosylmethionine decarboxylase proenzyme 1 (AMD1) had an up-regulation tendency in response to cold stress in the two samples. Moreover, the delta-1-pyrroline-5-carboxylate synthase (P5CS), δ-ornithine aminotransferase (δ-OAT), alanine-glyoxylate transaminase (AGXT), branched-chain-amino-acid transaminase (ilvE), alpha-aminoadipic semialdehyde synthase (AASS), Tyrosine aminotransferase (TAT) and arginine decarboxylase related to amino acid metabolism were identified in two cultivars variously expressed under cold stress. The above DEGs related to amino acid metabolism were suspected to the reason for amino acids content change. The RNA-seq data were validated by real-time quantitative RT-PCR of 19 randomly selected genes. The findings of our study provide the gene expression profile between two varieties of winter turnip rape, which lay the foundation for a deeper understanding of the highly complex regulatory mechanisms in plants during cold treatment.

Comparative proteomics analysis reveals the molecular mechanism of enhanced cold tolerance through ROS scavenging in winter rapeseed (Brassica napus L.).

Two winter rapeseed cultivars, "NS" (cold tolerant) and "NF" (cold sensitive), were used to reveal the morphological, physiological, and proteomic characteristics in leaves of plants after treatment at -4°C for 12 h(T1) and 24 h(T2), and at room temperature(T0), to understand the molecular mechanisms of cold tolerance. Antioxidant activity and osmotic adjustment ability were higher, and plasma membrane injury was less obvious, in NS than in NF under cold stress. We detected different abundant proteins (DAPs) related to cold tolerance in winter rapeseed through data-independent acquisition (DIA). Compared with NF, A total of 1,235 and 1,543 DAPs were identified in the NSs under T1 and T2, respectively. Compared with NF, 911 proteins were more abundant in NS only after cold treatment. Some of these proteins were related to ROS scavenging through four metabolic pathways: lysine degradation; phenylalanine, tyrosine, and tryptophan; flavonoid biosynthesis; and ubiquinone and other terpenoid-quinone biosynthesis. Analysis of these proteins in the four candidate pathways revealed that they were rapidly accumulated to quickly enhance ROS scavenging and improve the cold tolerance of NS. These proteins were noticeably more abundant during the early stage of cold stress, which was critical for avoiding ROS damage.

Screening and verification of reference genes for analysis of gene expression in winter rapeseed (Brassica rapa L.) under abiotic stress.

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 ß-actin and SAND in cold-stressed leaves, ß-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 PEG-stressed 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.

Transcriptome Profile Analysis of Winter Rapeseed (Brassica napus L.) in Response to Freezing Stress, Reveal Potentially Connected Events to Freezing Stress.

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.

Ecological and economic benefits of planting winter rapeseed (Brassica rapa L.) in the wind erosion area of northern China.

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.

Winter jujube (Zizyphus jujuba Mill.) quality forecasting method based on electronic nose.

Winter jujube (Zizyphus jujuba Mill.) quality forecasting method utilising electronic nose (EN) and double-layered cascaded series stochastic resonance (DCSSR) was investigated. EN responses to jujubes stored at room temperature were continuously measured for 8 days. Jujubes' physical/chemical indexes, such as firmness, colour, total soluble solids (TSS), and ascorbic acid (AA), were synchronously examined. Examination results indicated that jujubes were getting ripe during storage. EN measurement data was processed by stochastic resonance (SR) and DCSSR. SR and DCSSR output signal-to-noise ratio (SNR) maximums (SNR-MAX) discriminated jujubes under different storage time successfully. Multiple variable regression (MVR) results between physical/chemical indexes and SR/DCSSR eigen values demonstrated that DCSSR eigen values were more suitable for jujube quality determination. Quality forecasting model was developed using non-linear fitting regression of DCSSR eigen values. Validating experiments demonstrated that forecasting accuracy of this model is 97.35%. This method also presented other advantages including fast response, non-destructive, etc.