SummaryMultilocular silique is a desirable agricultural trait with great potential for the development of high‐yield varieties of Brassica. To date, no spontaneous or induced multilocular mutants have been reported in Brassica napus, which likely reflects its allotetraploid nature and the extremely low probability of the simultaneous random mutagenesis of multiple gene copies with functional redundancy. Here, we present evidence for the efficient knockout of rapeseed homologues of CLAVATA3 (CLV3) for a secreted peptide and its related receptors CLV1 and CLV2 in the CLV signalling pathway using the CRISPR/Cas9 system and achieved stable transmission of the mutations across three generations. Each BnCLV gene has two copies located in two subgenomes. The multilocular phenotype can be recovered only in knockout mutations of both copies of each BnCLV gene, illustrating that the simultaneous alteration of multiple gene copies by CRISPR/Cas9 mutagenesis has great potential in generating agronomically important mutations in rapeseed. The mutagenesis efficiency varied widely from 0% to 48.65% in T0 with different single‐guide RNAs (sgRNAs), indicating that the appropriate selection of the sgRNA is important for effectively generating indels in rapeseed. The double mutation of BnCLV3 produced more leaves and multilocular siliques with a significantly higher number of seeds per silique and a higher seed weight than the wild‐type and single mutant plants, potentially contributing to increased seed production. We also assessed the efficiency of the horizontal transfer of Cas9/gRNA cassettes by pollination. Our findings reveal the potential for plant breeding strategies to improve yield traits in currently cultivated rapeseed varieties.

a b s t r a c tAfforestation after deforestation using fast growing exotic species is creating major land use changes throughout China and the world. However, few studies have looked at changes in soil microbial communities resulting from the planting of exotic species. With paired comparison design, we studied the effects of replacing a native Pinus massoniana plantation with an exotic Eucalyptus (Eucalyptus urophylla  grandis) plantation on the composition and carbon metabolic function of soil microbial communities in Guangxi Province, southern China. We compared the microbial biomass, phospholipid fatty acid (PLFA) composition and carbon metabolic function (BIOLOG profiles) between both plantation types. Results showed the abundance of bacterial, fungal, actinomycetal, and total phospholipid fatty acids, and the proportion of 16:1x5c and five gram-negative characteristic bacterial PLFAs (of the six detected) in the Eucalyptus plantation soils were significantly lower than those of the P. massoniana plantations, as were biomass, carbon metabolic activity, and richness and diversity of the soil microbial community. The indicators denoting stress related to soil nutrient levels were significantly higher in the Eucalyptus plantation soils, such as the ratios of monounsaturated to saturated fatty acid, gram + to gram À bacteria, iso-to anteiso-branched PLFA, and cy19:0 to 18:1x7c. Canonical correspondence analysis (CCA) indicated the significant changes in the soil microbial community were mainly due to shrub and herb coverage, soil water content, soil organic carbon, soil N:P, and available N. Our research suggests forest conversion from native P. massoniana plantations to exotic Eucalyptus plantations alters the structure and function of the soil microbial community driven mainly by shifts in understory coverage and soil resource availability. Improved management practices, such as litter retention, reducing soil or understory disturbance during logging and subsequent establishment of the next rotation plantation, should be considered to help improve the metabolic function of soil microbial communities and increase soil resource availability during plantation management.

The oily wastewater generated in the industrial field is adversely affecting the environment, while the current methods for oil–water separation are complex and costly. Therefore, it is significant to use low cost and environmentally friendly materials to prepare a smart responsive superhydrophobic coating for the effective separation of oil–water mixtures. In this paper, a fluorine-free copolymer with pH responsiveness was fabricated by a solution impregnation method, and it was compounded by silica nanoparticles/polydimethylsiloxane to prepare a superhydrophobic coating on the paper and cotton fabric. The prepared superhydrophobic coating remained in the superhydrophobic state after the alkali treatment, while it would be converted into the hydrophilic state after the acid treatment. Therefore, the pH-responsive superhydrophobic coating will be applied in controlled selective oil–water separation.

Many studies have shown soil degradation after the conversion of native forests to exotic Eucalyptus plantations. However, few studies have investigated the long-term impacts of short-rotation forestry practices on soil microorganisms. The impacts of Eucalyptus successive rotations on soil microbial communities were evaluated by comparing phospholipid fatty acid (PLFA) abundances, compositions, and enzyme activities of native Pinus massoniana plantations and adjacent 1st, 2nd, 3rd, 4th generation Eucalyptus plantations. The conversion from P. massoniana to Eucalyptus plantations significantly decreased soil microbial community size and enzyme activities, and increased microbial physiological stress. However, the PLFA abundances formed "∪" shaped quadratic functions with Eucalyptus plantation age. Alternatively, physiological stress biomarkers, the ratios of monounsaturated to saturated fatty acid and Gram+ to Gram-bacteria, formed "∩" shaped quadratic functions, and the ratio of cy17:0 to 16:1ω7c decreased with plantation age. The activities of phenol oxidase, peroxidase, and acid phosphatase increased with Eucalyptus plantation age, while the cellobiohydrolase activity formed "∪" shaped quadratic functions. Soil N:P, alkaline hydrolytic nitrogen, soil organic carbon, and understory cover largely explained the variation in PLFA profiles while soil N:P, alkaline hydrolytic nitrogen, and understory cover explained most of the variability in enzyme activity. In conclusion, soil microbial structure and function under Eucalyptus plantations were strongly impacted by plantation age. Most of the changes could be explained by altered soil resource availability and understory cover associated with successive planting of Eucalyptus. Our results highlight the importance of plantation age for assessing the impacts of plantation conversion as well as the importance of reducing disturbance for plantation management.

Purpose Although it is generally accepted that planting exotic plant species alters metabolic function of soil microbial communities, its temporal dynamic is often ignored when evaluating ecological effects of associated land use changes. To investigate the dynamic impacts of successive Eucalyptus planting on carbon metabolic activities of soil microbial communities, we studied community-level physiological profiles of soil microbial communities in different generations of Eucalyptus plantations. Materials and methods We studied community-level physiological profiles of soil microbial communities, using the Biolog™ Ecoplates incubation, in adjacent first (G1), second (G2), third (G3), and fourth (G4) generation Eucalyptus plantations that were, respectively, aged 3, 8, 14, and 19 years in Guangxi province, southern China. We used the 'space-fortime substitution' approach to investigate the impact of stand age of exotic Eucalyptus plantations on carbon metabolic diversity and activities of soil microbial communities. For each Eucalyptus plantation generation, three experimental plots were randomly selected. In each plot, one composite soil sample from 0 to 10 cm in depth was obtained for the analyses. Results and discussion Single carbon source utilization varied with Eucalyptus plantation stand age. Among preselected 31 carbon sources, utilization of 17 carbon sources changed significantly, which was best described by a quadratic function (ten carbon sources) and an exponential function (seven carbon sources). As a result, cumulative averaged metabolic activity and metabolic diversity of soil microbial communities showed quadratic and exponential changes relative to Eucalyptus plantation stand age. The order of cumulative averaged carbon metabolic activity and metabolic diversity were G1>G4, G3>G2 and G1>G2>G3, G4 (p<0.05), respectively. The factors contributing to carbon source utilization structure of soil microbial communities for different stand ages of Eucalyptus plantations were shrub richness, soil organic carbon content, microbial biomass carbon, C-to-N ratio, and N-to-P ratio. Conclusions Eucalyptus plantation stand age has inconsistent non-linear impacts on two aspects of soil microbial metabolic function: (1) quadratic impacts on carbon metabolic efficiency and (2) exponential impacts on carbon metabolic diversity. The decreasing carbon metabolic diversity has no significant impact on carbon metabolic efficiency during successive Eucalyptus plantings. The results show that the importance of assessing long-term impacts of land use changes on soil microbial communities from exotic plantations by quantifying multi-aspect non-linear changes on soil microbial metabolic function.

The diversity of petal and leaf color can improve the ornamental value of rapeseed and promote the development of agriculture and tourism. The two copies of carotenoid isomerase gene (BnaCRTISO) in Brassica napus (BnaA09.CRTISO and BnaC08.CRTISO) was edited using the CRISPR/Cas9 system in the present study. The mutation phenotype of creamy white petals and yellowish leaves could be recovered only in targeted mutants of both BnaCRTISO functional copies, indicating that the redundant roles of BnaA09.CRTISO and BnaC08.CRTISO are vital for the regulation of petal and leaf color. The carotenoid content in the petals and leaves of the BnaCRTISO double mutant was significantly reduced. The chalcone content, a vital substance that makes up the yellow color, also decreased significantly in petals. Whereas, the contents of some carotenes (lycopene, α-carotene, γ-carotene) were increased significantly in petals. Further, transcriptome analysis showed that the targeted mutation of BnaCRTISO resulted in the significant down-regulation of important genes BnaPSY and BnaC4H in the carotenoid and flavonoid synthesis pathways, respectively; however, the expression of other genes related to carotenes and xanthophylls synthesis, such as BnaPDS3, BnaZEP, BnaBCH1 and BCH2, was up-regulated. This indicates that the molecular mechanism regulating petal color variation in B. napus is more complicated than those reported in Arabidopsis and other Brassica species. These results provide insight into the molecular mechanisms underlying flower color variation in rapeseed and provides valuable resources for rapeseed breeding.

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