Advances in Genome Editing With CRISPR Systems and Transformation Technologies for Plant DNA Manipulation

Satya Swathi Nadakuduti,Felix Enciso-Rodríguez

doi:10.3389/fpls.2020.637159

Abstract

The year 2020 marks a decade since the first gene-edited plants were generated using homing endonucleases and zinc finger nucleases. The advent of CRISPR/Cas9 for gene-editing in 2012 was a major science breakthrough that revolutionized both basic and applied research in various organisms including plants and consequently honored with “The Nobel Prize in Chemistry, 2020.” CRISPR technology is a rapidly evolving field and multiple CRISPR-Cas derived reagents collectively offer a wide range of applications for gene-editing and beyond. While most of these technological advances are successfully adopted in plants to advance functional genomics research and development of innovative crops, others await optimization. One of the biggest bottlenecks in plant gene-editing has been the delivery of gene-editing reagents, since genetic transformation methods are only established in a limited number of species. Recently, alternative methods of delivering CRISPR reagents to plants are being explored. This review mainly focuses on the most recent advances in plant gene-editing including (1) the current Cas effectors and Cas variants with a wide target range, reduced size and increased specificity along with tissue specific genome editing tool kit (2) cytosine, adenine, and glycosylase base editors that can precisely install all possible transition and transversion mutations in target sites (3) prime editing that can directly copy the desired edit into target DNA by search and replace method and (4) CRISPR delivery mechanisms for plant gene-editing that bypass tissue culture and regeneration procedures including de novo meristem induction, delivery using viral vectors and prospects of nanotechnology-based approaches.

Highlights

The year 2020 marks a decade since the first gene-edited plants were generated using homing endonucleases and zinc finger nucleases by traditional Agrobacterium mediated genetic transformation (Gao et al, 2010; Osakabe et al, 2010; Zhang et al, 2010)
The first CRISPR edited plants were developed in 2013 (Li et al, 2013; Shan et al, 2013) and since this technology has been applied in 45 plant genera across 24 families (Shan et al, 2020)
RNA VIRUSES AND MOBILE GUIDE RNAs FOR HERITABLE PLANT GENE-EDITING. Another heritable gene-editing method that has the potential of being a high-throughput method is by using a positive strand RNA virus, like the tobacco rattle virus (TRV) to deliver the single guide RNA (sgRNA) into Cas9 over-expressing plants via Agrobacterium infiltration (Ellison et al, 2020)

Summary

INTRODUCTION

The year 2020 marks a decade since the first gene-edited plants were generated using homing endonucleases and zinc finger nucleases by traditional Agrobacterium mediated genetic transformation (Gao et al, 2010; Osakabe et al, 2010; Zhang et al, 2010). Delivering Cas expression cassettes along with the sgRNA and growth regulators expressing cassettes via Agrobacterium into wild type plants is a feasible future approach which would facilitate DNA manipulation in a broad range of recalcitrant species Another heritable gene-editing method that has the potential of being a high-throughput method is by using a positive strand RNA virus, like the tobacco rattle virus (TRV) to deliver the sgRNAs into Cas over-expressing plants via Agrobacterium infiltration (Ellison et al, 2020). These modified sgRNAs are cloned into TRV vector which is delivered into plants by Agrobacterium infiltration (Figure 2B) All the mutations derived from M0 parents by this method were heritable (Ma et al, 2020)

NANOPARTICLES FOR DELIVERING BIOMOLECULES TO FACILITATE PLANT GENOME ENGINEERING

POTENTIAL FUTURE DEVELOPMENTS IN THE FIELD