Tuesday, 30 September 2014

Blogger's spell checker tries to fix "blog"

Why does blogger's spell check try to correct 'blog', and why does it suggest 'bog' or 'blag'. Are they telling me I'm writing s**t or trying to get something for free?

Please fix this one blogger.com.

Monday, 29 September 2014

Thanks for reading

This morning someone made the 500,000th page view on the CoreGenomics blog. It amazes me that so many people are reading this and the last couple of years writing have been really good fun. I've met many readers and some fellow bloggers, and received lots of feedback in the way of comments on posts, as well as at meetings. I've even had people recognise my name because of my blogging; surreal! But the last few years have seen some big changes in how we all use social media like blogs, Twitter, etc. I don't think there is a K-index for scientific bloggers, perhaps Neil can look at that one next ;-)

Question: What do you see?

Sunday, 28 September 2014

Making BaseSpace Apps in Bangalore

I'm speaking at the BaseSpace Apps developers conference in Bangalore tomorrow. It's my first App and my first time in India, so I'm pretty excited about the whole thing.

Tuesday, 23 September 2014

Welcome to a new company built around ctDNA analysis: Inivata

Inivata, is a new company spun out of Nitzan Rosenfelds research group at the CRUK Cambridge Institute (where I work). His group developed and published the TAm-seq method for circulating tumour DNA amplicon sequencing. The spin-out aims to develop blood tests measuring circulating tumour DNA (ctDNA) for use as a "liquid biopsy" in cancer treatment. Inivata has been funded  by Cancer Research Uk's technology arm CRT, Imperial Innovation, Cambridge Innovation Capital and Johnson & Johnson Development Corporation; initial funding has raised £4million.


Inivata is currently based in the Cambridge Institute and the start-up team include the developers of the TAm-seq method: Nitzan Rosenfeld (CRUK-CI), Tim Forshew (now at UCL Cancer Institute), James Brenton (CRUK-CI) and Davina Gale (CRUK-CI).

The research community has really taken hold of cell-free DNA and developed methods that are surpassing expectations. Cell-free DNA is having its largest impact outside of cancer in the pre-natal diagnostics market. And has been shown to be useful in many types of cancer. The use of ctDNA to follow tumour evolution was one of the best examples of what's possible I've seen so far and it's been exciting to be involved in some of this work. Inivata are poised to capitalise on the experience of the founding team and I'll certainly be following how they get on over the next couple of years.

If you fancy working in this field then they are currently hiring: molecular biologist, and computational biologist posts.

This is likely to become a crowded market as people pick up the tools available and deploy them in different settings. ctDNA is floating around in blood plasma and is ripe for analysis, I expect there is still lots of development space for new methods and ultimately I hope we'll be able to use ctDNA as a screening tool for early detection of cancer.

If we can enrich for mutant alleles using technologies like Boreal or Ice-Cold PCR then detection (not quantitation) may be possible far earlier than can be achieved today.

Monday, 15 September 2014

Are PCR-free exomes the answer

I'm continuing my exome posts with a quick observation. There have been several talks recently that I've seen where people present genome and exome data and highlight the drop-out of genomic regions primarily due to PCR amplification and hybridisation artefacts. They make a compelling case for avoiding PCR when possible, and for sequencing a genome to get the very best quality exome.

A flaw with this is that we often want to sequence an exome not simply to reduce the costs of sequencing, but more importantly to increase the coverage to a level that would not be economical for a genome, even on an X Ten! For studies of heterogeneous cancer we may want to sequence the exome to 100x or even 1000x coverage to look for rare mutant alleles. Unfortunately this is exactly the kind of analysis that might be messed up by those same PCR artefact's, namely PCR duplication (introducing allele bias) and base misincorporation (introducing artifactual variants).

PCR free exomes: In my lab we are running Illumina's rapid exomes so PCR is a requirement to complete the Nextera library prep. But if we were to use another method then in theory PCR-free exomes would be possible. Even if we stick to Nextera (or Agilent QXT) then we could aim for very low-cycle PCR libraries. The amount of exome library we are getting is huge, often 100's of nanomoles, when we only need picomoles for sequencing.

Something we might try testing is a PCR-free or PCR-lite (pardon the American spelling) exome to see if we can reduce exome artefacts and improve variant calling. If anyone else is doing this please let me know how you are getting along and how far we can push this.

Thursday, 4 September 2014

The newest sequencer on the block: Base4 goes public

I've heard lots of presentations about novel sequencing technologies, many have never arrived, some have come and gone, all have been pretty neat ideas; but so far not one has arrived that outperforms the Illumina systems many readers of this blog are using.

Base4's pyrophosphorolysis sequencing technology

The latest newcomer is Base4's single-molecule microdroplet sequencing technology. The picture above explains the process very well: a single molecule of double-stranded DNA is immobilised in the sequencer, single bases are cleaved at a defined rate from the 3' end by pyrophosphorolysis (the new Pyrosequencing perhaps?), as each nucleotide is cleaved it is captured into a microdroplet where it initiates a cascade reaction that generates a fluorescent signal unique to each base, as microdroplets are created at a faster rate than DNA is cleaved at the 3' end the system generates a series of droplets that can be read out by the sequencer (a little like the fluorescent products being read of a capillary electrophoresis instrument).

Base4 are talking big about what their technology can deliver. They say it will be capable of sequencing 1M bases per second with low systematic error rates. The single-molecules mean no amplification and read-lengths should be long. Parallelisation of the technology should allow multiple single-molecules to be sequenced at the same time. How much and when will have to wait a little longer.

I've been speaking to Base4 over the past few years after meeting their founder Cameron Frayling in a pub in Cambridge. Over the past two years Base4 has been developing their technology and recently achieved a significant milestone by demonstrating robust base-calling of single nucleotides in microdroplets. They are still small, with just 25 employees and are based outside Cambridge. I hope they'll be growing as we start to get our hands on the technology and see what it's capable of.

Low-diversity sequencing: RRBS made easy

Illumina recently released a new version of HCS v2.2.38 for the HiSeq. The update improves cluster definition significantly and enables low-diversity sequencing. It’s a great update and one that’s making a big impact on a couple of projects here.