Reproducibility of Illumina GoldenGate Assay clustering patterns. (A) Clustering patterns of the same SNP for WD (left) and FL (right) population can be different as shown in AA_scaffold1707_861. Cluster located in 0.0 position along x-axis is identified as heterozygous cluster in WD population; (B) Clustering patterns of the same SNPs in 96-plex (left) and 384-plex (right) assays are not highly reproducible. For AA_C444244_218 (top), the homozygous and heterozygous clusters are indistinguishable in 384-plex genotyping compared to 96-plex genotyping. A shift in heterozygous cluster position and lower signal intensities was observed in AA_scaffold17708_104 (bottom); (C) Two different SNPs from the same gene based on WD population can exhibit different clustering patterns although under the same influence of paralogous genes. In AA_scaffold17828 (top), SNP at position 1179 (left) generated a good cluster profile, but SNP at position 1028 (right) showed cluster compression. For gene AA_scaffold17912 (bottom), severe cluster compression was observed for SNP at position 330 (left) while the cluster located at 0.0 along x-axis were identified as heterozygote based on Mendelian inheritance in SNP at position 393 (right). Replicates of the parents from WD population are highlighted in yellow; Excluded samples are masked in grey color; The data points in color represent genotype calls for each sample (red = AA; purple = AB; blue = BB; black = unknown/outlier). The x-axis (Norm Theta) represents angle of the center of cluster in normalized polar coordinate while y-axis (Norm R) represent normalized intensity. In theory, AA, AB and BB clusters should have normalized theta (x-axis) values of 0.0, 0.5 and 1.0, respectively. Otherwise, the clusters exhibit cluster compression.
Wong et al. BMC Genomics 2012 13:726 doi:10.1186/1471-2164-13-726