Fig 1.
Scheme of experimental workflow developed in the current study.
Haploid nuclei were extracted from pollen grains, separated via flow-sorting and individually subjected to whole-genome-amplification (WGA). High quality samples, evaluated by PCR with chromosome 3H-specific primers, were genotyped using 25 KASP markers to measure crossover frequency and distribution.
Fig 2.
(A) Genotyping call rate of the positive controls across 24 KASP markers is indicated in red. Genomic DNA from Morex, Barke and Morex x Barke F1 plants was used. Two individual nuclei derived from Morex pollen grains were used to test for false allele calling due to whole-genome-amplification. The selected 50 Morex x Barke pollen nuclei samples are indicated in blue showing an average genotyping call rate of 71%. (B) Correlation between KASP genotyping call rate and preselection PCR call rate. r = 0.51, r² = 0.26.
Fig 3.
Comparison of the distribution of the number of crossovers.
(A) The relative frequency of the total number of crossovers per chromosome 3H grouped into classes ranging from 0 to 6 of the Morex x Barke pollen population (blue) in comparison to the Morex x Barke DH population data (red) [17]. (B) Correlation between KASP genotyping call rate and the number of crossovers found for each sample (r = -0.16, r² = 0.03).
Fig 4.
Recombination frequency along chromosome 3H determined by pollen genotyping.
(A) KASP marker positions (A to X) are shown as vertical bars along chromosome 3H. The physical length of the chromosome (Mbp) is shown on the x-axis. (B) The recombination frequency along chromosome 3H of a given physical interval measured as the proportion of crossovers to no-crossovers for each marker pair. A distal bias is shown by higher recombination frequencies towards the chromosome ends and low recombination frequencies between markers H and I.