Heritable CRISPR-Cas9 editing of plant genomes using RNA virus vectors

Abstract

Viral vectors hold enormous potential for genome editing in plants as transient delivery vehicles of CRISPR-Cas components. Here, we describe a protocol to assemble plant viral vectors for single guide RNA (sgRNA) delivery. The obtained viral constructs are based on compact T-DNA binary vectors of the pLX series and are delivered into Cas9-expressing plants through agroinoculation. This approach allows rapidly assessing sgRNA design for plant genome targeting, as well as the recovery of progeny with heritable mutations at targeted loci. For complete details on the use and execution of this protocol, please refer to Uranga et al.1 and Aragones et al.2. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=195 SRC="FIGDIR/small/507055v2_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@4a214corg.highwire.dtl.DTLVardef@d008d4org.highwire.dtl.DTLVardef@1f3b476org.highwire.dtl.DTLVardef@ccc792_HPS_FORMAT_FIGEXP M_FIG C_FIG