Optimising scalability
On average 30 targets per MinION run were amplified, pooled and
sequenced. Initially, a limited number of 11 samples were barcoded and
sequenced, but a reduction in usable data was noted (Supplementary Table
3). As such, this approach was replaced with careful sample pooling
instead. The 11 samples were not re-sequenced, as the small number meant
there was still ample data per target (mean of 19 million bases per
target). Targets were checked to ensure no overlap in genome location,
and demultiplexing was performed based on region extraction of the
mapped BAM files (see methods ‘Bioinformatics’). Using this approach, a
loss of 17.8% (SEM 7.2) of total basecalled yield was avoided by
bypassing barcode ligations, de-multiplexing binning issues and barcode
trimming (Supplementary Table 3).