CRISPR/Cas9-coupled recombineering for metabolic engineering of Corynebacterium glutamicum.

Genome engineering of Corynebacterium glutamicum, an important industrial microorganism for amino acids production, currently relies on random mutagenesis and inefficient double crossover events. Here we report a rapid genome engineering strategy to scarlessly knock out one or more genes in C. glutamicum in sequential and iterative manner. Recombinase RecT is used to incorporate synthetic single-stranded oligodeoxyribonucleotides into the genome and CRISPR/Cas9 to counter-select negative mutants. We completed the system by engineering the respective plasmids harboring CRISPR/Cas9 and RecT for efficient curing such that multiple gene targets can be done iteratively and final strains will be free of plasmids. To demonstrate the system, seven different mutants were constructed within two weeks to study the combinatorial deletion effects of three different genes on the production of γ-aminobutyric acid, an industrially relevant chemical of much interest. This genome engineering strategy will expedite metabolic engineering of C. glutamicum.

Recent Developments in Nanoporous Graphene Membranes for Organic Solvent Nanofiltration: A Short Review.

Graphene-based membranes are promising candidates for efficient organic solvent nanofiltration (OSN) processes because of their unique structural characteristics, such as mechanical/chemical stability and precise molecular sieving. Recently, to improve organic solvent permeance and selectivity, nanopores have been fabricated on graphene planes via chemical and physical methods. The nanopores serve as an additional channel for facilitating ultrafast solvent permeation while filtering organic molecules by size exclusion. This review summarizes the recent developments in nanoporous graphene (NG)-based membranes for OSN applications. The membranes are categorized depending on the membrane structure: single-layer NG, multilayer NG, and graphene-based composite membranes hybridized with other porous materials. Techniques for nanopore generation on graphene, as well as the challenges faced and the perspectives required for the commercialization of NG membranes, are also discussed.