Why should I sequence my SAGs at SCGC?

Specialized laboratory and computational procedures are essential in order to obtain high quality genomic sequences from SAGs. A major challenge to consider is the cross-talk of multiplexed Illumina libraries (also known as “sample bleeding” and “index switching”):

De novo assemblies of single amplified genomes (SAGs) are especially sensitive to this issue. This is due to the fact that single cell whole genome amplification is highly uneven across the genome, and deep sequencing is typically employed to facilitate the recovery of the under-amplified genome regions. As a result, even a relatively small overall fraction of mis-assigned reads may form sizeable contigs that represent over-amplified regions of co-sequenced SAGs. In 2014, SCGC established in-house infrastructure and procedures for the preparation of dual-barcoded libraries and their sequencing with NextSeq 500, followed by optimized data processing. These measures eliminated library cross-talk. To verify the efficacy of these solutions, SCGC benchmarks its entire workflow using SAGs of previously sequenced strains.

Please see the following links for additional information:

Strategies for achieving high sequencing accuracy for low diversity samples and avoiding sample bleeding using illumina platform
Index switching causing “spreading-of-signal” among multiplexed samples in illumina HiSeq 4000 DNA sequencing
The go-to gene sequencing machine with very strange results
Mixing sample types in a flowcell lane generated cross contamination artefacts
Index mis-assignment between samples on HiSeq 4000 and X-ten