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Related Experiment Video

Updated: Jun 27, 2025

Next-generation Sequencing of 16S Ribosomal RNA Gene Amplicons

Next-generation Sequencing of 16S Ribosomal RNA Gene Amplicons

Published on: August 29, 2014

ddRAD Sequencing and DNA Barcoding.

Vladislav Ivanov¹, Kyung Min Lee², Marko Mutanen¹

¹Ecology and Genetics Research Unit, University of Oulu, Oulu, Finland.

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

|April 29, 2024

Summary

This study details a double-digest restriction site-associated DNA sequencing protocol for capturing genomic single-nucleotide polymorphisms (SNPs). This method aids evolutionary studies, population structure inference, and species delimitation analyses.

Keywords:

Reduced genome representation methods Restriction sites SNPs

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Last Updated: Jun 27, 2025

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Published on: August 3, 2018

Area of Science:

Genomics
Molecular Biology
Bioinformatics

Background:

Restriction site-associated DNA sequencing (RADseq) is a powerful tool for population genomics.
Capturing single-nucleotide polymorphisms (SNPs) across genomes is crucial for evolutionary and ecological studies.
Existing RADseq protocols can be complex and require optimization.

Purpose of the Study:

To provide a detailed, step-by-step protocol for double-digest restriction site-associated DNA sequencing (ddRADseq).
To highlight essential equipment and common laboratory challenges associated with ddRADseq library preparation.
To facilitate the application of ddRADseq for various population genetic and phylogenetic analyses.

Main Methods:

Genomic DNA is chemically sheared using restriction enzymes.
Unique molecular tags are incorporated into DNA fragments.
Size selection is performed on the resulting DNA fragments.
Amplification of tagged DNA fragments is carried out.

Main Results:

The ddRADseq protocol effectively captures variable genomic sites, including SNPs.
The protocol is adaptable for diverse research questions in evolutionary biology.
Troubleshooting guidance for common laboratory issues is provided.

Conclusions:

The described ddRADseq protocol offers a robust method for generating genome-wide SNP data.
This protocol supports research in population structure, phylogenetics, gene flow, and species delimitation.
Standardized ddRADseq methods enhance the comparability of genomic data across studies.