Abstract

Increasing agricultural productivity via modern breeding strategies is of prime interest to attain global food security. An array of biotic and abiotic stressors affect productivity as well as the quality of crop plants, and it is a primary need to develop crops with improved adaptability, high productivity, and resilience against these biotic/abiotic stressors. Conventional approaches to genetic engineering involve tedious procedures. State-of-the-art OMICS approaches reinforced with next-generation sequencing and the latest developments in genome editing tools have paved the way for targeted mutagenesis, opening new horizons for precise genome engineering. Various genome editing tools such as transcription activator-like effector nucleases (TALENs), zinc-finger nucleases (ZFNs), and meganucleases (MNs) have enabled plant scientists to manipulate desired genes in crop plants. However, these approaches are expensive and laborious involving complex procedures for successful editing. Conversely, CRISPR/Cas9 is an entrancing, easy-to-design, cost-effective, and versatile tool for precise and efficient plant genome editing. In recent years, the CRISPR/Cas9 system has emerged as a powerful tool for targeted mutagenesis, including single base substitution, multiplex gene editing, gene knockouts, and regulation of gene transcription in plants. Thus, CRISPR/Cas9-based genome editing has demonstrated great potential for crop improvement but regulation of genome-edited crops is still in its infancy. Here, we extensively reviewed the availability of CRISPR/Cas9 genome editing tools for plant biotechnologists to target desired genes and its vast applications in crop breeding research.

Highlights

  • Food security is the most crucial challenge in the current scenario of a rapidly growing global population

  • Genome editing has emerged as a tremendous strategy for efficient and targeted genome manipulations, especially for crops which have complex genomes and which are difficult to improve through conventional breeding approaches [6]

  • We briefly describe first-generation genome editing tools such as transcription activator-like effector nucleases (TALENs), zinc-finger nucleases (ZFNs), and MNs and comprehensively elaborate on second-generation genome editing strategies with special focus on the applications of the CRISPR/Cas9 system in plant breeding for crop improvement

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Summary

Introduction

Food security is the most crucial challenge in the current scenario of a rapidly growing global population. The discovery of programmed sequence-specific nucleases (SSNs) has facilitated precise gene editing. In both plant and animal systems, application of SSNs for accurate GE has been recognized as a breakthrough in genome engineering. Non-homologous end joining is an error-prone DNA repair mechanism that facilitates direct end-joining of DSBs without involving a homologous template and can generate insertions or deletions at target sites to develop gene knockouts. In contrast to transgenic plants, genome-edited plants have the added benefit of site specificity [11] In breeding programs, these improved plants can be proven useful and subsequent species can be employed reliably with less concerns and comparatively minor monitoring methods are needed in contrast to traditional genetically engineered plants [12]

Modern Trends in Plant Genome Editing
TALENs
Construction of the sgRNA Expression Cassette
Construction of Cas9 Expression Cassettes
Strategies for Mutant Screening
Stable Expression
Transient Expression
Base Editing
Multiplex Genome Editing
Beyond Cas9
Regulatory Affairs of Genome-Edited Crops
Findings
Conclusion and Outlook

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