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Related Experiment Video

Updated: Jan 3, 2026

High-throughput CRISPR Vector Construction and Characterization of DNA Modifications by Generation of Tomato Hairy Roots

High-throughput CRISPR Vector Construction and Characterization of DNA Modifications by Generation of Tomato Hairy Roots

Published on: April 30, 2016

CRISPR-Cas-Mediated Gene Knockout in Tomato.

Gwen Swinnen^1,2, Thomas Jacobs^1,2, Laurens Pauwels^1,2

¹Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, Belgium.

Methods in Molecular Biology (Clifton, N.J.)

|November 21, 2019

Summary

CRISPR-Cas genome editing enables the creation of stable tomato gene knockout lines. This method facilitates plant functional genomics by generating loss-of-function mutants for specific genes.

Keywords:

CRISRP-Cas Gene knockout Genome editing Loss-of-function mutation Null mutation Site-directed mutagenesis Solanaceae Solanum lycopersicum Targeted mutagenesis Tomato

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Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format

Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format

Published on: April 8, 2017

Area of Science:

Plant Genomics
Molecular Biology
Biotechnology

Background:

Loss-of-function mutants are essential for understanding gene function in plant functional genomics.
CRISPR-Cas genome editing technology has advanced the ability to generate targeted mutations in various plant species, including tomato (Solanum lycopersicum).

Purpose of the Study:

To describe a methodology for utilizing CRISPR-Cas genome editing to generate stable gene knockout lines in tomato.
To provide a framework for creating null alleles for specific genes in tomato for functional studies.

Main Methods:

Employing CRISPR-Cas9 system for targeted DNA double-strand break (DSB) induction at specific genomic loci in tomato.
Leveraging the nonhomologous end joining (NHEJ) repair pathway to introduce small insertions or deletions (indels) resulting in gene knockouts.
Selection and verification of stable knockout lines through appropriate screening methods.

Main Results:

Successful generation of targeted gene knockouts in tomato using CRISPR-Cas genome editing.
Demonstration of the efficiency of CRISPR-Cas in creating null alleles through NHEJ-mediated repair.
Establishment of stable tomato gene knockout lines suitable for further functional genomics research.

Conclusions:

CRISPR-Cas genome editing is a powerful and accessible tool for generating stable tomato knockout lines.
This approach significantly aids in advancing plant functional genomics research by enabling precise gene function analysis.
The described protocol provides a reliable method for creating loss-of-function mutants in tomato.