Sequence capture across large phylogenetic scales by using pooled PCR-generated baits: A case study of Lepidoptera

Area of Science:

• Evolutionary Biology
• Genomics
• Phylogenetics

Background:

• Hybridization capture for large-scale phylogenetics is limited by the genetic distance between bait and target DNA.
• Existing methods face challenges in efficiently capturing genetic data across diverse evolutionary lineages.

Purpose of the Study:

• To develop a novel strategy for preparing PCR-generated baits to overcome limitations in sequence capture across large phylogenetic scales.
• To create a universal set of nuclear markers for Lepidoptera (moths and butterflies) to facilitate phylogenetic analysis.
• To demonstrate the effectiveness of pooled DNA from representative species for bait preparation.

Main Methods:

• A DNA pool was created from six representative Lepidoptera species spanning six superfamilies.
• This pooled DNA was used to generate PCR baits for 94 nuclear protein-coding genes.
• The homemade PCR baits were employed for sequence capture in 43 species across 17 Lepidoptera families.

Main Results:

• The strategy successfully recovered 94% of the targeted nuclear loci across the sampled Lepidoptera species.
• Phylogenetic trees constructed from two datasets (~90 kb and ~120 kb) showed high similarity and resolution, with 90% of nodes exhibiting >95% bootstrap support.
• The method demonstrated reliable capture of numerous nuclear markers across diverse Lepidoptera lineages.

Conclusions:

• Pooling DNA from representative species is an effective strategy for preparing PCR baits to minimize genetic distance for sequence capture.
• The newly developed set of 94 nuclear markers provides a valuable phylogenetic tool for Lepidoptera research.
• This PCR bait preparation strategy can accelerate data collection and advance the application of sequence capture techniques in phylogenetics.