Some comments and analysis from the exciting and fast moving world of Genomics. This blog focusses on next-generation sequencing and microarray technologies, although it is likely to go off on tangents from time-to-time
Exome sequencing (@Exome_seq) can be used to generate CNV maps bit the data are
noisy compared to WGS or SNP-arrays. In this post I’ll describe a novel workflow
to get high-quality CNV from an exome-seq pipeline and show some very new data…
Image reproduced and enlarged at the bottom of this post.
Back to the paper: The MD Anderson group used their gene panel on FFPE material and reported on the "sensitivity, specificity, reproducibility, and applicability of using the Ion Proton 409-gene panel to routinely screen for SNVs, insertions/deletions, and CNVs". To do this they used 55 tumours (20 with paired normals) and four cell lines.
Fig 6A: ERBB2 amplifications as seen in 4 breast cancer samples.
The Ion Torrent Ampliseq Comprehensive Cancer Panel uses 4000 primer pairs across four primer pools each requiring just 15ng on input DNA (60ng total), and up to 96 samples can be indexed using Ion Xpress Barcodes. Alignment and analysis of data was performed using the Torrent Suite software. And the groups OncoSeek was used to generate a clinical report.
Concordance between platforms was high when the group compared results to an earlier PGM panel of 46 genes, and detected
pretty much everything they expected. The Proton did call some InDels
that the PGM missed, which the group put down to improvements in calling
The group reported high sensitivity for SNVs up to 5% allelic fraction, and high reproducibility. This, coupled to the minimal FFPE input, and fast turnaround (5 days) makes the platform combination one that could be used in a clinical molecular diagnostics laboratory. Additionally they reported that up to 10 samples could be multiplexed per run and that this fitted in particularly well with their workflows.
There is lots of suggestion that longevity has a genetic component, but I can't help but consider that the environmental component is likely to be stronger, and this would mask the genetics. Kim's study was very small, just 17 genomes, so the chances of finding anything were equally small. But had there been low-hanging fruit this would almost certainly have been a Science paper rather than PLoS One (sorry PLoS).
In the Summer I posted a Christmas letter to Santa. He's already delivered number 2 (cheaper RNA-seq) and 3 (longer RNA-seq reads via PE250), and number 4 might be coming soon (exomes at PE125). I'd also asked that he not deliver HiSeq X Ten as an individual instrument just yet, but as there are just 44 days left till Christmas I thought I'd look head and see what might be the reasons for, or not for gift wrapping a single X Ten this Christmas.
I was chatting with a colleague at work who'd asked me if I know anywhere they could get their genome or exome sequenced. My genome has been sat in the freezer for over five years wanting to go onto a flow cell, but I've never been comfortable putting it on our own machines. I did get 23andMe'd a few years ago but they've closed the exome for now.
Today there are many sequencing service providers across the world. Would any of them be open to a consumer led project? How many genomes/exomes would we need to sequence to get a price consumers were willing to pay? To test the market we've used AllSeq: "the global sequencing marketplace", and a couple of replies have now come in!
What Casey has done is pretty simple and it is very well explained in his blog post, or by Rob Lanfear who has posted instructions on GitHub. There are three simple steps for PubMed and Twitter (and more for arxiv, peerj, etc).
This was not a planned post but it follows on nicely from today's other one about exomes. This time I'm writing about Fluidigm's new single-cell exome-seq protocol. Yup that's right, whole exomes from 96 single cells! The C1 is an amazing piece of kit (wish I had one) and we've used it a little bit for mRNA-seq. The ability to sequence single-cell genomes and exomes means you can pretty much do whatever you want with a single-cell now. So how do the exomes look?