RFP peptide

Escherichia coliBL21 is arguably the most popular host for industrial production of proteins, and industrial fermentations are often plagued by phage infections. The CRISPR/Cas system is guided by a gRNA to cleave a specific DNA cassette, which can be developed into a highly efficient programable phage defense system. In this work, we constructed a CRISPR/Cas system targeting multiple positions on the genome of T7 phage and found that the system increased the BL21's defense ability against phage infection. Furthermore, the targeted loci on phage genome played a critical role. For better control of expression of CRISPR/Cas9, various modes were tested, and the OD of the optimized strain BL21(pT7cas9, pT7-3gRNA, prfp) after 4 h of phage infection was significantly improved, reaching 2.0, which was similar to the control culture without phage infection. Although at later time points, the defensive ability of CRISPR/Cas9 systems were not as obvious as that at early time points. The viable cell count of the engineered strain in the presence of phage was only one order of magnitude lower than that of the strain with no infection, which further demonstrated the effectiveness of the CRISPR/Cas9 phage defense system. Finally, the engineered BL21 strain under phage attack expressed RFP protein at about 60% of the un-infected control, which was significantly higher than the parent BL21. In this work, we successfully constructed a programable CRISPR/Cas9 system to increase the ability ofE. coliBL21's to defend against phage infection, and created a resistant protein expression host. This work provides a simple and feasible strategy for protecting industrialE. colistrains against phage infection.

The generation of burst pulses in a multi-pass passively mode-locked laser is systematically explored. We exploit intracavity mirrors with different thicknesses to serve as reflected Fabry-Perot (RFP) cavities in laser resonators. Using a RFP cavity with the thinnest optical thickness, we can generate picosecond optical burst pulses with the highest intra-burst pulse rate of 33.12 GHz. The maximum output power of 2.6 W is obtained at an incident pump power of 11.9 W. We further develop an analytical model which considers the RFP effects to manifest the first-order autocorrelation and spectral behaviors in experiments. The agreement between theoretical analyses and experimental explorations provides the feasibility to flexibly manipulate the burst modes in practical applications. (C) 2019 Optical Society of America