Figure 1
From: Massively parallel sequencing for early molecular diagnosis in Leber congenital amaurosis
Workflow overview. For each patient, all exons and intron–exon boundaries of the 16 known LCA genes are amplified using qPCR, followed by random ligation and shearing. Subsequently, 12 amplicon pools are indexed and sequenced in one lane of the Illumina Genome Analyzer IIx. (a) Cumulative distribution plot of mean Cq values of both replicates for each patient of lane 1. (b) Cumulative distribution plot of mean end-point fluorescence values of both replicates for each patient of lane 1. (c) Merged Agilent Bioanalyzer electropherograms (DNA 7500) of purified pooled PCR-product (blue, 1/4 dilution), purified end-repaired DNA (green, 1/4 dilution), and ligated DNA (red, 1/3 dilution) from patient 7. (d) Agilent Bioanalyzer electropherogram (DNA 1000) of sheared product from patient 7. (e) Screenshot of the NextGENe alignment viewer showing the homozygous AIPL1 mutation c.885del, identified in patient 17. Cq, quantification cycle; LCA, Leber congenital amaurosis; qPCR, quantitative PCR.
