The enzymes and the sizes of the restriction fragments are optimized with training data to ensure even distribution and avoid repeats. The fragments are also selected over a tight range, to optimize the PCR reaction. The protocol is similar to ddRAD, for example with a first digestion with MseI, heat inactivation and a second digestion with AluI. The resulting fragments are PCR amplified, adaptors added and the fragments purified.

Pros:
Deep sequencing for genotyping accuracy
Reduced representation strategy to reduce sequencing costs
Pre-designed reduced representation scheme to optimize marker efficiency
Double barcode system for large populations
Cons:
Does not cover whole genome

MseI

MseI, an isoschizomer of RspRSII, is a restriction enzyme that cleaves within the recognition sequence 5'-TTAA-3'.

AluI