CRISPR/Cas: A powerful tool for gene function study and crop improvement

Abstract

Background: It is a long-standing goal of scientists and breeders to precisely control a gene for studying its function as well as improving crop yield, quality, and tolerance to various environmental stresses. The discovery and modification of CRISPR/Cas system, a nature-occurred gene editing tool, opens an era for studying gene function and precision crop breeding.

Aim of review: In this review, we first introduce the brief history of CRISPR/Cas discovery followed the mechanism and application of CRISPR/Cas system on gene function study and crop improvement. Currently, CRISPR/Cas genome editing has been becoming a mature cutting-edge biotechnological tool for crop improvement that already used in many different traits in crops, including pathogen resistance, abiotic tolerance, plant development and morphology and even secondary metabolism and fiber development. Finally, we point out the major issues associating with CRISPR/Cas system and the future research directions.Key Scientific Concepts of Review: CRISPR/Cas9 system is a robust and powerful biotechnological tool for targeting an individual DNA and RNA sequence in the genome. It can be used to target a sequence for gene knockin, knockout and replacement as well as monitoring and regulating gene expression at the genome and epigenome levels by binding a specific sequence. Agrobacterium-mediated method is still the major and efficient method for delivering CRISPR/Cas regents into targeted plant cells. However, other delivery methods, such as virus-mediated method, have been developed and enhanced the application potentials of CRISPR/Cas9-based crop improvement. PAM requirement offers the CRISPR/Cas9-targted genetic loci and also limits the application of CRISPR/Cas9. Discovering new Cas proteins and modifying current Cas enzymes play an important role in CRISPR/Cas9-based genome editing. Developing a better CRISPR/Cas9 system, including the delivery system and the methods eliminating off-target effects, and finding key/master genes for controlling crop growth and development is two major directions for CRISPR/Cas9-based crop improvement.

Keywords: CRISPR; Cas; Crop improvement; Gene function; Genome editing; PAM.

Graphical abstract

Fig. 1

Application of CRISPR/Cas genome editing in gene functional study. CRISPR/Cas system has a diversity of application in gene functional study. Based the DNA double strand break (DSB) repair mechanism, CRISPR can directly cause gene knockout (silencing) by insertion or deletion of a couple of nucleotides and repaired by non-homologous end join (EHEJ); however, if the homologue-directed repair (HDR) happed, with a DNA donor, CRISPR/Cas genome editing can be used to replace an undesirable gene or over express (knockin) and an individual gene. If deactivating the Cas9 enzyme, and with transcription effector or other enzymes fused with the dCas9, CRISPR/Cas system also can be used to base editing, epigenome editing and imaging.

Fig. 2

Application of CRISPR/Cas genome editing in crop improvement, CRISPR/Cas is a robust tool and have huge potentials on crop improvement. Though targeting an individual DNA sequence, CRISPR/Cas can be used to add and/or change the expression of an individual gene that controls plant tolerance to abiotic and biotic stresses as well as plant growth and development. CRISPR/Cas also can change the secondary metabolism for controlling the biosynthesis of protein, carbohydrate, oil, and functional components. Partial figures are modified from previous publications , .