2b or not 2b? 2bRAD is an effective alternative to ddRAD for phylogenomics

Area of Science:

• Genomics
• Evolutionary Biology
• Bioinformatics

Background:

• Restriction-site-associated DNA sequencing (RADseq) is crucial for generating genome-wide single-nucleotide polymorphisms (SNPs) for phylogenetic inference.
• Empirical comparisons of different RADseq protocols (ddRAD and 2bRAD) are lacking, hindering informed method selection.

Purpose of the Study:

• To empirically evaluate how ddRAD and 2bRAD methods impact phylogenetic estimation in non-model frogs.
• To compare these methods regarding phylogenetic information, missing data, allelic dropout, and sequencing depth effects.
• To assess laboratory effort, computational time, costs, and repeatability for each method.

Main Methods:

• Phylogenetic analysis of two non-model frog groups using ddRAD and 2bRAD data.
• Comparison of phylogenetic signal, missing data, and allelic dropout across methods and sequencing depths.
• Evaluation of practical aspects: lab work, computation, cost, and repeatability.

Main Results:

• Both ddRAD and 2bRAD yielded well-supported phylogenetic trees, even at low sequencing depths, with comparable missing data and phylogenetic signal.
• 2bRAD demonstrated higher repeatability, simpler lab protocols, and faster bioinformatics assembly compared to ddRAD.
• Fewer parsimony-informative sites were obtained from 2bRAD data using native pipelines, indicating a need for pipeline optimization.

Conclusions:

• Both ddRAD and 2bRAD are viable RADseq methods for phylogenetic studies, offering distinct advantages and disadvantages.
• 2bRAD is recommended for its efficiency and repeatability, provided downstream analyses can accommodate potentially fewer informative sites.
• Empirical comparisons are vital for selecting the most appropriate RADseq method based on specific research needs and resources.