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Abstract: As the mechanism of the soil-fungi -plant interaction under the tobacco intercropping model is unclear, and the contribution of this triple interaction to tobacco plant growth is still difficult to predict, so, in this paper, Illumina high-throughput sequencing technology was used to analyze the effects of monoculture (CK) and intercropping with garlic (T1 and T2 treatments) on rhizosphere soil nutrients, enzyme activities, fungal community, tobacco plant growth, and to evaluate the regulation of rhizosphere soil microenvironment and plant productivity on intercropping. The results showed that intercropping significantly increased the nutrient, enzyme activity, fungal community diversity of rhizosphere soil and tobacco plant biomass. Although the dominant fungi in each treatment were the same at the gate level, the distribution ratio of dominant fungi was different. Correlation analysis showed that Ascomycota was positively correlated with all the nutrient and enzyme activity indexes, and the nutrient and enzyme activity indexes were positively correlated with the tobacco plants growth, which strongly indicated that the soil microenvironment under intercropping had a potential effect on the production performance of tobacco plants. In conclusion, tobacco intercropping not only increased the plants biomass, significantly improved the nutrients and enzyme activities of rhizosphere soil, optimized the composition and diversity of fungal community, which may be the result of soil-fungi-plant interaction.Abstract: As the mechanism of the soil-fungi -plant interaction under the tobacco intercropping model is unclear, and the contribution of this triple interaction to tobacco plant growth is still difficult to predict, so, in this paper, Illumina high-throughput sequencing technology was used to analyze the effects of monoculture (CK) and intercropping with ga...Learn More
Abstract: By analyzing the hairpin structure of miR-486, it was found that the mature sequences of miR-486-5p and miR-486-3p are complementary. For this reason, it is probably the greatest challenge to express miR-486-5p using a lentiviral expression vector in C2C12 cells. Thus, we constructed shRNA/siRNA lentiviral expression vectors for microRNA-486-5p based on miR-30/451 and miRNA sponges. Oligonucleotide sequences (miR-486-5p TuD (tough decoy RNAs), miR-486-5p sponge, miR-486-5p-shRNA30 and miR-486-5p-shRNA451) were digested and cloned into the PLV-U6-MCS vector to generate U6-driven expression cassettes for mmu-miR-486-5p, and these vectors, including anti-miR-486-5p TuD, anti-miR-486-5p sponge, miR-486-5p-shRNA30 and miR-486-5p-shRNA451, were assessed by sequencing. The lentivirus vector was packaged in 293T cells according to the manufacturer’s instructions. Subsequently, C2C12 cells were infected using a lentivirus for the expression of miR-486-5p, and this lentivirus of PLV-U6-MCS served as a control. qRT-PCR analysis showed that anti-miR-486-5p TuD and the anti-miR-486-5p sponge decreased the expression level of miR-486-5p in C2C12 cells by 0.5-fold and 0.7-fold, respectively. Additionally, miR-486-5p-shRNA30 and miR-486-5p-shRNA451 upregulated the expression level of miR-486-5p in C2C12 cells by 15-fold and 3-fold, respectively. However, the expression level of miR-486-3p was increased up to 1000-fold in C2C12 cells containing the miR-486-5p-shRNA30 lentivirus, while other lentiviruses only led to 3-fold increases compared with the miR-486–5p-shRNA30 lentivirus in C2C12 cells. Additionally, western blot and gray density analysis demonstrated that the expression levels of AKT, and pAKT, ERK1/2 and pERK1/2, and Snail were upregulated (approximately 1.20-fold to 1.33-fold and 1.05-fold to 1.20-fold, respectively) in C2C12 cells infected with the anti-miR-486-5p TuD lentivirus and anti-miR-486-5p sponge lentivirus and downregulated (approximately 0.63–0.96 times and 0.72–0.92 times, respectively) in C2C12 cells infected with the miR-486-5p-shRNA30 lentivirus and miR-486-5p-shRNA451 lentivirus. In summary, anti-miR-486-5p TuD, which dramatically reduced the expression level of miR-486-5p and upregulated endogenous protein levels in C2C12 cells, was superior to the anti-miR-486-5p sponge in terms of interference expression of miR-486-5p. miR-486-5p-shRNA451, which did not lead to substantial expression of miR-486-3p and remarkably downregulated endogenous protein levels in C2C12 cells, was better than miR-486-5p-shRNA30 for overexpression of miR-486-5p. These results suggested that anti-miR-486-5p TuD and miR-486-5p-shRNA451 provide a simple way to build long-term effective overexpression or interference of miR-486-5p, to study miRNA function in C2C12 and mice, and to create a potential diagnostic method based on materials for diseases caused by miRNA deregulation.Abstract: By analyzing the hairpin structure of miR-486, it was found that the mature sequences of miR-486-5p and miR-486-3p are complementary. For this reason, it is probably the greatest challenge to express miR-486-5p using a lentiviral expression vector in C2C12 cells. Thus, we constructed shRNA/siRNA lentiviral expression vectors for microRNA-486-5p bas...Learn More
