70 research outputs found
2-kW Average Power CW Phase-Conjugate Solid-State Laser
We have demonstrated stable operation of a 2-kW Yb:YAG phase-conjugate master oscillator power amplifier (PC-MOPA) laser system with a loop phase-conjugate mirror (LPCM). This is the first demonstration of a continuous wave (CW)-input LPCM MOPA operating at a power greater than 1 kW with a nearly diffraction-limited output beam. The single-pass beam quality incident on the LPCM varied with the specific operating conditions, but it was typically times diffraction-limited (XDL). The measured beam quality with an MOPA output power of 1.65 kW was 1.3 XDL
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Kinetic characterization of CRISPR-Cas12a DNA targeting
Cas12a is a type V class 2 CRISPR-Cas nuclease that has recently been repurposed as a precision DNA-cutting tool, alongside the type II Cas9. Their potential for biotechnological and genome editing applications has driven enthusiastic efforts to biochemically characterize their assembly and RNA-guided DNA targeting mechanisms. Two important obstacles that must be resolved before these nucleases can be used widespread are their specificity and efficiency; CRISPR nucleases are known to cut sequences that resemble their intended target, and within the cell—where our genomic DNA is packaged into protein-rich chromatin—it is unclear how target preferences are influenced. Here, we quantitatively study Acidaminococcus sp. Cas12a DNA targeting behavior in vitro using a kinetics-based approach. We begin by characterizing Cas12a binding and cleavage on matched and mismatch targets (Chapter 2), uncovering how Cas12a achieves such high levels of specificity despite cleaving DNA in a rate-limiting binding regime. We then characterize Cas12a DNA targeting on substrates that begin resemble genomic chromatin (Chapter 3). Lastly, we review the fundamentals of CRISPR nuclease specificity and how engineering efforts to develop a ‘high-fidelity’ nuclease should be focused (Chapter 4).Cellular and Molecular Biolog
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