Lactuca georgica Grossh. is a wild species belonging to the secondary lettuce gene pool: additional evidence, obtained by KASP genotyping

This work evaluated the genetic relationships among 442 single-seed descent accessions, representing eight Lactuca spp., including five wild Lactuca relatives (WLRs) (Lactuca georgica, L. altaica, L. saligna, L. serriola, L. aculeata), L. tuberosa, L. undulata, and the domesticated lettuce, L. sativa, 437 of them representing a core subset of the Institute of Evolution's Wild Lettuce Gene Bank (IoE's WLGB) collection. The analysis was performed by profiling 115 single-nucleotide polymorphism markers with the fluorescent KASP genotyping assay. KASP marker fragments were scored as either allele “A” or allele “B”, and then treated as bi-allelic data, but the dataset also included a relatively high number of U-scores, noting the absence of the specific sequence, that were treated as missing data. Often U-scores were specific for a certain species. Data analysis of the five WLRs showed that allele frequencies of 103 (97.2%) out of 106 differentiating loci varied significantly among the species, where 59.7% of the KASP marker diversity was between species. A neighbor-network analysis involving samples of the five WLRs and a single L. sativa cv. clearly clustered all 430 samples in accordance with their taxonomic determination. These results obtained from multiple complementary analyses of large natural populations and individuals, question the assignment of L. georgica to the primary lettuce gene pool. Together with our previous results obtained by using TRAP markers and hybridization experiments, we conclude that L. georgica is a constituent of the secondary lettuce gene pool. Our analyses shed also light on the proper placement of L. altaica.