Figure 1.

Workflow to generate and analyse long‐read amplicons in pools. (a) Leaf material is collected using a size template to ensure equal sample representation in each pool. Long‐range amplicon products are obtained by PCR with direct barcoding to individually tag each pool. Products are visualized by gel electrophoresis for quality control and validation of amplicon concentration measurements. (b) All population pools are combined in equal amounts in a single tube. A PacBio library is generated and sequenced in circular consensus mode on a Sequel II system. (c) Computational processing includes read‐consensus building, demultiplexing and filtering of raw reads. (d) pbaa clustering is used for variant detection and filtering (Kronenberg et al., 2021). The output is fasta files listing all haplotypes per population pool and including meta information on read coverage of each haplotype.