Gene editing in plants: assessing the variables through a simplified case study

Authors: Shockey, Jay

Submitted to: Plant Molecular Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/31/2020
Publication Date: 2/10/2020
Citation: Shockey, J. 2020. Gene editing in plants: assessing the variables through a simplified case study. Plant Molecular Biology. 103:75-89. https://doi.org/10.1007/s11103-020-00976-2.
DOI: https://doi.org/10.1007/s11103-020-00976-2

Interpretive Summary: Successful engineering of plants for production of biobased chemical feedstocks and other valuable components is often hindered by the ability to guide metabolism towards the desired pathways, and away from undesirable pathways. This usually occurs due to the activity of genes that researchers would like to switch off, and replace with genes with desirable properties. Recent developments have created the ability to target specific genes for deactivation through a process often called gene editing or genome editing, but much more basic knowledge is necessary to learn exactly how to harness this power effectively. Criteria for successful gene editing were studied and contrasted here, and the results may be useful to help other labs, just starting out with their own gene editing plans, to bypass some of the bottlenecks to success.

Technical Abstract: The recent advent of genome editing technologies (especially CRISPR, clustered regularly interspaced short palindromic repeats) has revolutionized various fields of scientific research. The process is much more specific than previous mutagenic processes and allows for targeting of nearly any gene of interest for the creation of loss-of-function mutations and many other types of editing, including gene-replacement and gene activation. However, not all CRISPR construct designs are successful, due to several factors, including differences in the strength and cell- or tissue-type specificity of the regulatory elements used to express the nuclease and single-guide RNA components, and differences in the relative editing efficiency at different target areas within a given gene. Here we compare the levels of successful heritable editing created in Arabidopsis thaliana by constructs containing different combinations of promoters and single guide RNA sequences. Significant differences in gene editing levels were observed using both specific and non-specific targeting approaches, suggesting that all elements of CRISPR construct design must be carefully considered.