Formerly at Corteva Agriscience™, DuPont Knowledge Centre, Hyderabad, Telangana, India.
Corteva Agriscience , Johnston, IA-50131, USA.
Methods Mol Biol. 2021;2238:115-134. doi: 10.1007/978-1-0716-1068-8_8.
CRISPR-Cas resonates a revolutionary genome editing technology applicable through a horizon spreading across microbial organism to higher plant and animal. This technology can be harnessed with ease to understand the basic genetics of a living system by altering sequence of individual genes and characterizing their functions. The precision of this technology is unparallel. It allows very precise and targeted base pair level edits in the genome. Here, in the current chapter, we have provided end-to-end process outline on how to generate genome edited plants in crops like rice to evaluate for agronomic traits associated with yield, disease resistance and abiotic stress tolerance, etc. Genome editing process includes designing of gene editing strategy, vector construction, plant transformation, molecular screening, and phenotyping under control environment conditions. Furthermore, its application for development of commercial crop product may require additional processes, including field trials in the target geography for evaluation of product efficacy. Evaluation of genome edited lines in controlled greenhouse/net house or open field condition requires few generations for outcrossing with wild-type parent to eliminate and/or reduce any potential pleiotropic effect in the edited genome which may arise during the process. The genome edited plant selected for advancement shall harbor the genome with only the intended changes, which can be analyzed by various molecular techniques, advanced sequencing methods, and genomic data analysis tools. CRISPR-Cas-based genome editing has opened a plethora of opportunities in agriculture as well as human health.
CRISPR-Cas 引发了一场革命性的基因组编辑技术革命，其应用范围横跨微生物到高等植物和动物。这项技术可以轻松地用于通过改变单个基因的序列并描述其功能来了解生命系统的基本遗传学。这项技术的精确性是无与伦比的。它允许在基因组中进行非常精确和靶向的碱基对水平编辑。在当前这一章中，我们提供了一个端到端的流程概述，介绍如何在水稻等作物中生成经过基因组编辑的植物，以评估与产量、抗病性和抗非生物胁迫等相关的农艺性状。基因组编辑过程包括设计基因编辑策略、载体构建、植物转化、分子筛选以及在控制环境条件下进行表型分析。此外，其在商业作物产品开发中的应用可能需要额外的过程，包括在目标地理区域进行田间试验，以评估产品的功效。在受控温室/网室或开放田间条件下评估基因组编辑系需要与野生型亲本进行几代的杂交，以消除和/或减少编辑基因组中可能在过程中产生的任何潜在的多效性影响。选择用于推进的基因组编辑植物应仅具有预期的变化的基因组，这可以通过各种分子技术、先进的测序方法和基因组数据分析工具进行分析。基于 CRISPR-Cas 的基因组编辑为农业和人类健康带来了众多机遇。
