High-Throughput and Low-Cost Genotyping Method for Plant Genome Editing

Liu, Lei, Chen, Richelle, Fugina, Christopher John, Siegel, Ben, Jackson, David (April 2021) High-Throughput and Low-Cost Genotyping Method for Plant Genome Editing. Curr Protoc, 1 (4). e100. ISSN 2691-1299

URL: https://www.ncbi.nlm.nih.gov/pubmed/33826801
DOI: 10.1002/cpz1.100

Abstract

Genome editing technologies have revolutionized genetic studies in the life sciences community in recent years. The application of these technologies allows researchers to conveniently generate mutations in almost any gene of interest. This is very useful for species such as maize that have complex genomes and lack comprehensive mutant collections. With the improvement of genome editing tools and transformation methods, these technologies are also widely used to assist breeding research and implementation in maize. However, the detection and genotyping of genomic edits rely on low-throughput, high-cost methods, such as traditional agarose gel electrophoresis and Sanger sequencing. This article describes a method to barcode the target regions of genomic edits from many individuals by low-cost polymerase chain reaction (PCR) amplification. It also employs next-generation sequencing (NGS) to genotype the genome-edited plants at high throughput and low cost. This protocol can be used for initial screening of genomic edits as well as derived population genotyping on a small or large scale, at high efficiency and low cost. © 2021 Wiley Periodicals LLC. Basic Protocol 1: A fast genomic DNA preparation method from genome edited plants Basic Protocol 2: Barcoding the amplicons of edited regions from each individual by two rounds of PCR Basic Protocol 3: Bioinformatics analysis.

Item Type: Paper
Subjects: organism description > plant > maize
Investigative techniques and equipment > assays > next generation sequencing
CSHL Authors:
Communities: CSHL labs > Jackson lab
SWORD Depositor: CSHL Elements
Depositing User: CSHL Elements
Date: April 2021
Date Deposited: 30 Apr 2021 19:46
Last Modified: 30 Apr 2021 19:46
Related URLs:
URI: https://repository.cshl.edu/id/eprint/39964

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