Genome-Wide Analysis and Functional Characterization of LACS Gene Family Associated with Lipid Synthesis in Cotton (Gossypium spp.).

Cotton (Gossypium spp.) is the fifth largest oil crop in the world, and cottonseed provides abundant vegetable oil resources and industrial bioenergy fuels for people; therefore, it is of practical significance to increase the oil content of cotton seeds for improving the oil yield and economic benefits of planting cotton. Long-chain acyl-coenzyme A (CoA) synthetase (LACS) capable of catalyzing the formation of acyl-CoAs from free fatty acids has been proven to significantly participate in lipid metabolism, of which whole-genome identification and functional characterization of the gene family have not yet been comprehensively analyzed in cotton. In this study, a total of sixty-five LACS genes were confirmed in two diploid and two tetraploid Gossypium species, which were divided into six subgroups based on phylogenetic relationships with twenty-one other plants. An analysis of protein motif and genomic organizations displayed structural and functional conservation within the same group but diverged among the different group. Gene duplication relationship analysis illustrates the LACS gene family in large scale expansion through WGDs/segmental duplications. The overall Ka/Ks ratio indicated the intense purifying selection of LACS genes in four cotton species during evolution. The LACS genes promoter elements contain numerous light response cis-elements associated with fatty acids synthesis and catabolism. In addition, the expression of almost all GhLACS genes in high seed oil were higher compared to those in low seed oil. We proposed LACS gene models and shed light on their functional roles in lipid metabolism, demonstrating their engineering potential for modulating TAG synthesis in cotton, and the genetic engineering of cottonseed oil provides a theoretical basis.

Multiplex real-time SYBR Green I PCR assays for simultaneous detection of 15 common enteric pathogens in stool samples.

Diarrheal diseases account for more than 50% of foodborne diseases worldwide, the majority of which occur in infants and young children. The traditional bacterial detection method is complex and time-consuming; therefore, it is necessary to establish a rapid and convenient detection method that can detect multiple pathogens simultaneously. In this study, we developed a set of five multiplex real-time SYBR Green I PCR assays to simultaneously detect 15 common enteric pathogens based on the Homo-Tag Assisted Non-Dimer system. These assays effectively reduced primer-dimer formation and improved the stability, uniformity, and amplification efficiency of multiplex PCR. The detection limit of the multiplex SYBR Green I PCR system was approximately 104-106 CFU/mL for stool specimens. Furthermore, we vitrified heat-unstable components on the cap of a reaction tube, showing that Taq DNA polymerase, dNTPs, primers, and SYBR Green I remained stable at 25 °C. In summary, we developed multiplex SYBR Green I PCR assays that can simultaneously detect 15 enteric pathogens. This method is comprehensive, rapid, inexpensive, accurate, and simple and displays high specificity.

Inhibition of migration and invasion by berberine via inactivation of PI3K/Akt and p38 in human retinoblastoma cell line.

BACKGROUND: As a clinically important natural isoquinoline alkaloid, berberine has been reported to possess various pharmacological effects. OBJECTIVES: This study was aimed to investigate the effect of berberine on cell migration and invasion in human retinoblastoma (Rb) cells. MATERIAL AND METHODS: The cytotoxicity of berberine was estimated by 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay. After being stimulated with berberine under various concentrations, the cell migration and invasion were evaluated by transwell assay. Then, the expression levels of epithelial-mesenchymal transition (EMT) markers were determined by quantitative reverse transcription PCR (qRT-PCR) and western blot analysis. Furthermore, the phosphorylation levels of protein kinase B (Akt) and p38 were detected by western blot analysis. Finally, the effect of phosphatidylinositol-3-kinase (PI3K) and p38 inhibitors on cell migration and invasion was estimated by transwell assay. Untreated cells acted as control for all the experiments. RESULTS: The concentrations of berberine for further studies were controlled in a range of 0 to 100 µM. The cell migration and invasion were both suppressed by berberine in a dose-dependent manner compared to the control (p < 0.05 or p < 0.001). Berberine remarkably down-regulated expression of E-cadherin and up-regulated expression of vimentin and α-SMA compared to the control (p < 0.01 or p < 0.001). Furthermore, the phosphorylation levels of Akt and p38 were both down-regulated by berberine in comparison to the control. Furthermore, the addition of berberine accompanied by LY294002 or SB203580 significantly suppressed cell migration and invasion compared to the addition of berberine alone (p < 0.05). CONCLUSIONS: Berberine suppressed cell migration and invasion via inactivation of PI3K/Akt and p38.