RADseq as a valuable tool for plants with large genomes-A case study in cycads

Area of Science:

• Genomics
• Plant Science
• Evolutionary Biology

Background:

• Full genome sequencing is challenging and expensive for organisms with large, complex genomes, such as cycads (Cycadales), which possess genomes up to 60 Gbp.
• Restriction site-associated DNA sequencing (RADseq) is a cost-effective method for identifying genome-wide markers in various organisms.

Purpose of the Study:

• To evaluate the effectiveness of RADseq, specifically the ezRAD protocol, for generating genome-wide markers across all 10 genera of Cycadales.
• To adapt and apply RADseq to cycads, overcoming challenges posed by their large genomes and high plastid numbers.

Main Methods:

• Applied ezRAD protocol to samples from all 10 Cycadales genera, including a specific dataset for Cycas calcicola (72 samples).
• Utilized existing plastid and mitochondrial genomes for read mapping to recover genomic regions.
• Performed de novo assembly to identify phylogenetically informative nuclear loci.

Main Results:

• Successfully recovered plastid, mitochondrial, and nuclear markers across all cycad genera.
• De novo assembly yielded up to 138,407 high-depth clusters and 1,705 informative loci per genus, with 4,421 loci for C. calcicola.
• Marker numbers were sufficient for downstream analyses and could be increased by adjusting assembly parameters.

Conclusions:

• Demonstrated the successful application of RADseq for generating a substantial number of genome-wide markers in cycads, despite their large genomes.
• The modified RADseq protocol is suitable for cycads and other organisms with large genomes, providing a valuable tool for genetic and evolutionary studies.