Fast gene disruption in Trichoderma reesei using in vitro assembled Cas9/gRNA complex

CRISPR/Cas9 has wide application potentials in a variety of biological species including Trichoderma reesei, a filamentous fungus workhorse for cellulase production. However, expression of Cas9 heterologously in the host cell could be time-consuming and sometimes even troublesome. We tested two gene disruption methods in T. reesei using CRISPR/Cas9 in this study. The intracellularly expressed Cas9 led to unexpected off-target gene disruption in...