Computational Tools and Resources for CRISPR/Cas Genome Editing.

The past decade has witnessed a rapid evolution in identifying more versatile clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) nucleases and their functional variants, as well as in developing precise CRISPR/Cas-derived genome editors. The programmable and robust features of the genome editors provide an effective RNA-guided platform for fundamental life science research and subsequent applications in diverse scenarios, including biomedical innovation and targeted crop improvement. One of the most essential principles is to guide alterations in genomic sequences or genes in the intended manner without undesired off-target impacts, which strongly depends on the efficiency and specificity of single guide RNA (sgRNA)-directed recognition of targeted DNA sequences. Recent advances in empirical scoring algorithms and machine learning models have facilitated sgRNA design and off-target prediction. In this review, we first briefly introduce the different features of CRISPR/Cas tools that should be taken into consideration to achieve specific purposes. Secondly, we focus on the computer-assisted tools and resources that are widely used in designing sgRNAs and analyzing CRISPR/Cas-induced on- and off-target mutations. Thirdly, we provide insights into the limitations of available computational tools that would help researchers of this field for further optimization. Lastly, we suggest a simple but effective workflow for choosing and applying web-based resources and tools for CRISPR/Cas genome editing.

Functional Differentiation of BnVTE4 Gene Homologous Copies in α-Tocopherol Biosynthesis Revealed by CRISPR/Cas9 Editing.

Tocopherols are essential nutrients for human health known as vitamin E. Vitamin E deficiency can have a profound effect on human health, including the central nervous system and cardiovascular and immune protection. Multiple enzymatic steps are involved in the conversion between different forms of tocopherols. Among them, γ-tocopherol methyltransferase encoded by gene VTE4 catalyzes the conversion of γ- to α-tocopherol or δ- to ß-tocopherol isoforms. However, the gene copies and their functional contribution of VTE4 homologs in Brassica napus were not elucidated. To this end, different mutation combinations of four putative BnVTE4 homologous copies were generated by using CRISPR/Cas9 genome editing technology. Editing of those BnVTE4 homologs led to a significant change of the α-tocopherol content and the ratio between α- and γ-tocopherol compared with wide-type control. Analysis of the different combinations of BnVTE4-edited homologs revealed that the contribution of the BnVTE4 individual gene displayed obvious functional differentiation in α-tocopherol biosynthesis. Their contribution could be in order of VTE4.C02-2 (BnaC02G0331100ZS) > VTE4.A02-1 (BnaA02G0247300ZS) > VTE4.A02-2 (BnaA02G0154300ZS). Moreover, the VTE4.A02-1 and VTE4.A02-2 copies might have severe functional redundancies in α-tocopherol biosynthesis. Overall, this study systemically studied the different effects of BnVTE4 homologs, which provided a theoretical basis for breeding high α-tocopherol content oilseed rape.

Dynamic Interference Behaviors of Arbitrary Multilayer Commingling Production in Heavy Oil Reservoirs with Water Flooding.

To obtain sustainable economical oil production and recovery of investment, some oil fields adopted the strategy of multilayer commingling production at an early stage. This leads to interlayer interference and losing part of the recoverable reserves. In this paper, dynamic interference behaviors of arbitrary multilayer commingling production in heavy oil reservoirs are analyzed. Based on the non-Darcy flow equation, the Buckly-Leverett equation, and the material balance equation, a mathematical model of arbitrary multilayer commingling production is obtained. Oil and water relative permeability, saturation, and bottom hole flow pressure microelement and the iteration method are employed to solve the mathematical model in the time domain. The new model is verified by comparing the results from the typical black oil model using the Darcy law. The sensitivity analysis of critical parameters on interference behaviors, such as permeability, oil viscosity, effective drainage boundary, and voidage replacement ratio, is carried out. The model obtained in this paper can be used for oil and liquid productivity analysis during the overall process of commingling production and extended to be applied in numerical experiments with different combinations of typical parameters as well.

Expression of microRNA-210 and the prognosis in glioma patients: a meta-analysis.

Aim: To evaluate the predictive power of tumor microRNA-210 (miR-210) on overall survival (OS) in glioma patients. Materials & methods: Studies were identified through searching PubMed, Embase and China National Knowledge Internet electronic databases. Meta-analyses were performed with a random- or fixed-effect model according to the heterogeneity. Results: Six studies were included. Results showed that increased miR-210 expression in tumor independently predicted poor OS in glioma patients (hazard ratio [HR]: 1.38; p = 0.001). Subgroup analyses showed that the prognostic efficacy of tumor miR-210 levels for OS was stronger in overall patients with glioma (HR: 2.22; p < 0.001) than in those with glioblastoma (HR: 1.13; p = 0.01). Conclusion: Expression of miR-210 may predict poor survival in patients with glioma.