Targeted Base Editing in Soybean Using a CRISPR-Cas9 Cytidine Deaminase Fusion

Abstract

AbstractBase editing has emerged as a novel and efficient genome editing approach for helping agronomic traits improvement. This editing system can revert a single-base change or SNP without double-strand break in the genome. Cytosine base editors (CBEs) and adenine base editors (ABEs) are two types of base editors, which are composed of a deactivated Cas9 endonuclease (dCas9) or a Cas9 nickase (nCas9) and a catalytic domain which is capable of deaminating a cytosine or adenine, respectively. CBEs convert C·G to T·A. In our lab, we have constructed a CBE editor for soybean base editing. The CBE is composed of nCas9, a rat cytidine deaminase enzyme APOBEC1, and a uracil DNA glycosylase inhibitor (UGI). The CBE was constructed in the PTF101 vector background. The PTF101-CBE has successfully achieved single-base alteration in soybean. Here, we present a detailed protocol of this base editor to generate a point mutation in soybean.Key words Soybean Base editingCytosine base editors Cas9 nickase Point mutation