Home cooks know well that putting vegetables in the crisper can keep them healthy; new work shows that putting CRISPR in the vegetables works as well. Every year, plant viruses destroy tens of billions of dollars worth of crops around the world. Even worse, localized crop failures can cause food shortages and destroy family farmers. Viruses are particularly troublesome because there are no sprays that target them directly.
Fortunately, the scientifically red-hot CRISPR-cas system of genome editing fame actually has its roots in a bacterial defense system against viruses. In fact, CRISPR has even been used to defend human blood cells against HIV. Now, Xiang Ji and Huawei Zhang, working in Caixia Gao’s lab in Beijing, have planted the Cas9 nuclease into wild mustard and tobacco relatives, showing that the system works in the vegetable kingdom as well.
Screening for Good Cas9 Target Sites
First, the team constructed a library of plasmid vectors containing cas9 as well as guide RNAs (gRNAs) targeting a variety of loci in a test geminivirus. Next, they transfected these vectors into N. benthamiana leaves by agroinoculation (using Agrobacterium to transfer the DNA). After waiting two days, they infected the same leaves with a geminivirus, in particular, beet severe curly top virus (BSCTV), to test for immunity.
Encouragingly, some of the gRNAs protected the leaves from BSCTV symptoms, and the CRISPR system also prevented the virus from replicating and spreading, as assayed by qPCR on leaf tissue. As is its wont, CRISPR induced small indels in the remaining viral genomes, with most mutations at the target site being short, 1-10 bp deletions.
Next, the team made transgenic N. benthamiana and Arabidopsis plants stably expressing cas9 and a high-efficiency gRNA. Just like the transiently transfected leaves, whole plants expressing the CRISPR system protected themselves from the geminivirus. Different plants expressed cas9 at different levels, and those expressing it more highly were better at preventing virus replication.
So don’t let yourself succumb to a severe curly top! Check out the full paper available from Nature Plants, 2015.