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- Targeted single cell DNA sequencing without prior whole genome amplification for mutational analysis of circulating tumor cells

# Targeted single cell DNA sequencing without prior whole genome amplification for mutational analysis of circulating tumor cells

Single-cell DNA sequencing usually begins with whole genome amplification, whose bias drops real mutations and adds false ones; this AACR 2019 poster from RareCyte and the University of Washington reports what happened when circulating tumor cells were sequenced straight from cell lysate instead.

Presented by RareCyte and University of Washington.

Read or download to see:

- Skipping whole genome amplification raised median read depth 7-fold and reduced target dropout. Cells pulled from the WGA protocol at 4 successive steps showed amplification bias arising as early as the pre-amplification step and increasing throughout the run; libraries built straight from cell lysate instead covered the amplicon panel more evenly.

- Non-WGA libraries recovered 12 of 15 known A549 variants, against 8 of 15 after WGA. Mean variant-detection sensitivity rose from 51% to 77%, and specificity moved the same way: the false-positive error frequency of non-WGA samples was under 5% of the WGA samples, which the authors attribute to polymerase errors introduced during WGA itself.

- In blood from a prostate cancer patient, the method recovered both mutations cell-free DNA had already flagged. Guardant360 cfDNA sequencing identified somatic TP53 and PTEN mutations; targeted sequencing of individual CTCs retrieved with CytePicker detected both in 4 of 5 CTCs and in 0 of 3 white blood cell controls, with TP53 G244S reads at that locus ranging from 44% to 100%.

[Download poster](https://cdn.sanity.io/files/s3p6uqps/production/d53952701d68ce95a558c1b2cbd6591dcf69613f.pdf)

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[Download the poster](https://cdn.sanity.io/files/s3p6uqps/production/d53952701d68ce95a558c1b2cbd6591dcf69613f.pdf) to view it.

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