Table 2.
Library preparation | Sequencing platform | Max CAG repeat size | Max modal CAG size * | Max somatic expansion size | Estimated sequencing cost per sample ** | Quantity of genomic DNA required per sample | Indels | Relative accuracy of the quantification of somatic expansions | Relative accuracy of the quantification of somatic contractions |
Bulk-PCR | Illumina MiSeq | ∼115 | ∼90 | ∼120 | ∼$8 † | 20 ng | No | + | not detected if |
PacBio RSII | ≥550 | ∼250 | ≥550 | ∼$50 † | 0.1 to 1.2 μg | Yes | + | not very large | |
PacBio Sequel | ≥550SM | ∼250SM | ≥550SM | ∼$30 † | 20 to 240 ngSM | Yes | + | ||
PacBio Sequel II | ≥550SM | ∼250SM | ≥550SM | ∼$10 † | 20 to 60 ngSM | Yes | + | ||
SP-PCR | Illumina MiSeq | ∼115SM | ∼90SM | ∼120SM | ∼$8 ‡ | 2 ngSLM | No | ++ | + |
PacBio RSII | ≥1500L | ≥1200L | ≥1500L | ∼$50 ‡ | 10 to 120 ngSLM | Yes | ++ | + | |
PacBio Sequel | ≥1500LM | ≥1200LM | ≥1500LM | ∼$30 ‡ | 2 to 24 ngSLM | Yes | ++ | + | |
PacBio Sequel II | ≥1500LM | ≥1200LM | ≥1500LM | ∼$10 ‡ | 2 to 6 ngSLM | Yes | ++ | + | |
Amplification- | Illumina MiSeq | ∼115S | ∼90S | ∼120S | ∼$20 † | ≥5μgL | No | +++ | +++ |
free | PacBio RSII | ≥1500L | ≥150L | ≥1500L | ∼$1000 † | ≥5μgL | Yes | +++ | +++ |
PacBio Sequel | ≥1500LM | ≥150LM | ≥1500LM | ∼$600 † | ≥5μgL | Yes | +++ | +++ | |
PacBio Sequel II | ≥1500LM | ≥150LM | ≥1500LM | ∼$100 † | ≥5μgL | Yes | +++ | +++ | |
Amplicon | Illumina MiSeq | ∼115S | ∼90S | ∼120S | ∼$20 # | 20 to 150 ngL | No | +++ | +++ |
sequencing | PacBio RSII | ≥550SM | ∼250SM | ≥550SM | ∼$1000 # | 2.5 to 30μgSL | Yes | +++ | +++ |
of barcoded | PacBio Sequel | ≥550SM | ∼250SM | ≥550SM | ∼$600 # | 0.4 to 4.8μgSLM | Yes | +++ | +++ |
single molecules | PacBio Sequel II | ≥550SM | ∼250SM | ≥550SM | ∼$100 # | 0.2 to 0.6μgSLM | Yes | +++ | +++ |
Observations described in this study are indicated in bold, the other information corresponds to expectations for approaches not used in this study based on observations described in this study (S), on what has been described on other trinucleotide loci in the literature (L) and/or on the manufacturer information available for each sequencing platform (M). *: Max modal allele size for which the modal allele size can be estimated by sequencing and for which somatic mosaicism will reliably be quantified. **: based on [63]. †: assuming a minimum of ∼5,000 reads per sample and a maximum of 384 samples per sequencing run. ‡: assuming 20 SP-PCRs per sample and 250 reads per SP-PCRs. #: assuming ∼20 reads per single molecule and the genotyping of 5,000 single molecules.