Table 1 Performance of cLPP target capture and sequencing
From: Multiplex target capture with double-stranded DNA probes
(i) Sensitivity; percentage of the target bases that are represented by one or more reads | 98.7% of all exons and 97.8% of all target bases at >20× coverage (0.012× mean coverage) |
(ii) Specificity; percentage of sequence reads that map to the intended targets (5,471 exons) | 98.1% mapped target reads confirming the high on-target specificity of LPP capture |
(iii) Accuracy; base calling concordance to known sample SNVs | >99% concordance rate for both heterozygous and homozygous SNVs with coverage of 97.9%; sample SNVs at >20× coverage (0.012× mean) |
(iv) Uniformity; variability in sequence coverage across target regions | 91% of capture products were distributed within a 50-fold range (94% within 100-fold) |
(v) Reproducibility; or how closely results obtained from independent samples correlate | r = 0.93 rank-order correlation between two different HapMap samples (Figure S8 in Additional file 1); 1.15% SD (P = 0.01) for detecting heterozygous SNV (>20×) in five cLPP capture experiments. |
(vi) Cost of LPP target capture | $86 per sample for 100 genes in 100 samples or $16.70 per sample for 100 genes in 1,000 samples (Figure S2 in Additional file 1). |
(vii) Ease of use and time effort | <8 hours target capture and library preparation ('sequencing-ready'), <24 hours MiSeq and approximately 6 hours variant calling (524 genes). Total time: 38 hours |
(viii) DNA amount required per experiment | >50 ng of genomic DNA |
(ix) Multiplexing of candidate genomic targets and of DNA samples | Multiplex target capture of 524 genes per sample; sample multiplexing of 7 capture libraries per MiSeq run (#B, Additional file 2). |