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Related Experiment Video

Updated: Aug 9, 2025

Author Spotlight: Streamlining Rice Breeding with CRISPR/Cas for Obtaining Optimal Phenotypic and Agronomic Traits

Author Spotlight: Streamlining Rice Breeding with CRISPR/Cas for Obtaining Optimal Phenotypic and Agronomic Traits

Published on: January 3, 2025

Precise and heritable gene targeting in rice using a sequential transformation strategy.

Wenxin Zhang^1,2, Rui Wang^1,2, Dali Kong^1,2

¹Shanghai Center for Plant Stress Biology, CAS Center for Excellence in Molecular Plant Sciences, Chinese Academy of Sciences, Shanghai 200032, China.

Cell Reports Methods

|February 23, 2023

Summary

This summary is machine-generated.

This study introduces a sequential transformation strategy for precise gene targeting in rice. The CRISPR-Cas9 system significantly enhances gene knockin efficiency, aiding rice research and breeding.

Keywords:

CRISPR-Cas9 DNA methylation OsFTL1 OsROS1a epigenetics gene targeting genome editing genome engineering rice

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Published on: February 14, 2020

Area of Science:

Plant molecular biology
Genetics and genomics
Biotechnology

Background:

Gene targeting (GT) is crucial for modifying plant genomes, but its efficiency is low in higher plants.
Engineered sequence-specific nucleases (SSNs) can enhance GT frequency by inducing double-strand breaks (DSBs).
A CRISPR-Cas9-mediated sequential transformation strategy was previously reported for GT in Arabidopsis.

Purpose of the Study:

To establish an efficient gene targeting method in rice using the sequential transformation strategy.
To improve the efficiency of heritable gene targeting in the Japonica rice cultivar Oryza sativa Nipponbare.
To demonstrate precise gene knockin into specific endogenous loci in rice.

Main Methods:

Generated two independent parental lines expressing maize Ubiquitin 1 promoter-driven Cas9.
Utilized Agrobacterium-mediated transformation to deliver sgRNA and donor DNA.
Applied the sequential transformation strategy for CRISPR-Cas9-mediated gene targeting.

Main Results:

Achieved efficient sequential transformation-mediated gene targeting in Oryza sativa Nipponbare.
Successfully performed precise GFP (Green Fluorescent Protein) knockin into the OsFTL1 and OsROS1a loci.
Demonstrated the effectiveness of strong Cas9 activity and robust DSBs for GT establishment.

Conclusions:

The developed CRISPR-Cas9 sequential transformation method enables efficient and precise gene targeting in rice.
This GT technology holds significant potential for applications in rice research and crop improvement.
The strategy facilitates targeted genomic modifications, advancing rice breeding programs.