Single molecule genomics

Lead Research Organisation: MRC Laboratory of Molecular Biology

Abstract

My group works on ways to analyse DNA the genetic material. Most methods analyse large DNA samples, which have to be produced by cloning or other methods. instead, we concentrate on analysing single DNA molecules. We have developed a range of techniques with this theme. Recently, we have devised a way to look for copy-number changes: mutations or natural variations which lead to extra copies or missing copies of some pieces of DNA. This method has proved valuable in analysing the changes that happen in the genome during the development of cancer. |We are also developing microfluidic tools: miniaturized analytical devices based around glass chips containing minute tubes, mixing chambers and other scaled-down versions of laboratory equipment. With these devices, we intend to streamline and automate the analysis of copy-number changes. We are also working on ways to sequence (that is, to read) large pieces of DNA quickly, again using microfluidic tools.

Technical Summary

Our focus is on developing and applying new methods of analysing the genome. Our approaches revolve around the analysis of single DNA molecules, for which we have developed a range of techniques.|Single-molecule methods simplify many aspects of genome analysis. Not only is it unnecessary to purify a genomic fragment by cloning (since a single molecule is always pure), but we can readily access aspects of genomic variation which are masked by analysing bulk samples. |Recently, we developed a method for accurately measuring copy-number variations which arise either in cancer (for example, as a result of non-reciprocal translocations or regional genomic amplification), or as part of the normal spectrum of variation. In essence, we just count the number of molecules of a given target sequence in very dilute DNA samples. The approach is robust and sensitive, and can be applied to even minute amounts of material. Ongoing work as shown that we can use this method to obtain otherwise inaccessible information on the genomic changes that happen in the earliest stages of lung cancer. Other, collaborative projects are focussed on breast and colorectal cancers, and other groups have also adopted the method.|We are also developing microfluidic tools for genomic analysis. One of these, a single-chip copy-number device, will greatly streamline and accelerate our existing method for measuring copy-number, allowing a compact device to give accurate and rapid copy-number measurements in minutes. A second device, whose development will take longer, will enable us to sequence large (>>10kb) fragments of genomic DNA rapidly and cheaply, in contrast to traditional Sanger sequencing or to sequence tag methods such as 454 and Solexa.

Publications

10 25 50
 
Description Research strategies in Malaysia
Geographic Reach Europe 
Policy Influence Type Participation in advisory committee
Impact Since 2007, I have been a visiting professor for 1-2 weeks per year in Malaysia. This has involved reviews of their research programs and discussions with their Science Minister and with senior members (deputy VC and below) at one of their leading research universities (UKM). I am trying to foster a more solid base for science in Malaysia which, as a developing nation in which research is still emerging, has the opportunity to follow a more sustainable and productive course than other countries (Singapore, Brazil) which have attempted to become significant players in molecular biology and genomics. During my latest visit (2011), I have given workshops on research strategy and seminars on the research ethos at LMB. One of my focusses has been to caution them against over-bureaucratization, and to point out that LMB's success over the years has been in spite of, rather than because of, the current Western fashion for evaluation, accountability and quantifiable short-term deliverables.
 
Title Microfluidics infrastructure 
Description I have established a microfluidics fabrication facility at LMB. Use of this facility has been scaled down following the recent quinquennial review, but it will be of use. 
Type Of Material Improvements to research infrastructure 
Year Produced 2011 
Provided To Others? Yes  
Impact Sequencing platform developed with the aid of this facility is now the subject of commercial exploitation. 
 
Title Molecular Copy-number counting 
Description MCC is a technique which allows analysis of copy-number changes (gains or losses of DNA) in samples which are not tractable to other methods. It is particularly valuable in the accurate, multi-locus analysis of cancer biopsies. 
Type Of Material Technology assay or reagent 
Year Produced 2007 
Provided To Others? Yes  
Impact See publications. In my own group, we have used this method to identify an important gene whose amplification is INVARIABLY associated with the development of one common type of lung cancer, and which is a promising target for diagnosis, prognosis and intervention. In collaboration with Daser (University of Mainz), we have developed a method for analysing aneuploidy (uniquely, at the level of individual CHROMATIDS) in human polar bodies, leading to a method of pre-fertilisation diagnosis of oocyte aneuploidy; trials of this method in patients are underway. Patents have been filed for MCC (granted), for a combined assay which couples MCC with multilocus mutational analysis (under way) and for the adaptation of MCC to pre-fertilisation diagnosis (under way) 
 
Title SOX2 as a key gene in lung cancer 
Description We identified SOX2 as a critical gene in the transition from low-grade dysplasia to pre-cancerous high-grade dysplasia in lung cancer. SOX2 is being further investigated as a potential therapeutic target. 
Type Of Material Model of mechanisms or symptoms - human 
Provided To Others? No  
Impact Ongoing studies on SOX2 
 
Description Analysis of ancient DNA 
Organisation Max Planck Society
Country Germany 
Sector Charity/Non Profit 
PI Contribution Development and implementation of methods for analysing DNA in ancient specimens
Collaborator Contribution Developed techniques for analysing multiple sequences from fossil specimens (mammoth, Neanderthal)
Impact Publications. More generally, our methods are now commonly used in the analysis of ancient DNA.
Start Year 2007
 
Description Detection and identification of human pathogens and parasites 
Organisation Public Health England
Country United Kingdom 
Sector Public 
PI Contribution Shortly before being made redundant by the MRC, I devised a sensitive diagnostic for neonatal meningitis. This is now being trialled at Addenbrooke's and in Cardiff.
Collaborator Contribution Yes
Impact Yes
Start Year 2014
 
Description Development of novel sequencing technology 
Organisation Base4 Innovation
Country United Kingdom 
Sector Private 
PI Contribution Since my redundancy from MRC in April 2015, I have continued to develop this technology, which is now the sole activity of a 40-strong biotech company in Cambridge.
Collaborator Contribution Base4 has increased its commitment to this project, with approx. 40 employees full-time.
Impact Yes
Start Year 2012
 
Description Pre-fertilisation diagnosis 
Organisation SH-Gen
Country Germany 
Sector Private 
PI Contribution Since my redundancy from MRC in 2015, this work has led to clinical applications; it is now being offered to patients at one of Germany's leading fertility centres, and is proving uniquely effective in improving the pregnancy rates from IVF.
Collaborator Contribution Yes
Impact Currently in use. The main outputs are babies.
Start Year 2006
 
Title Molecular Copy-number counting 
Description Method for analysing changes in copy-number in DNA 
IP Reference ES2337207 
Protection Patent granted
Year Protection Granted 2010
Licensed No
Impact Identification of driver genes in colon and lung cancer.
 
Title New sequencing technology. 
Description I invented a new sequencing technology. A contract has been signed with Cambridge-based company Base4 for the joint development of this technology; this contract covers mutual confidentiality agreements and the terms for handling IP, which is being filed jointly by Base4 and MRC. 
IP Reference  
Protection Protection not required
Year Protection Granted 2012
Licensed Commercial In Confidence
Impact Main impact is that I am currently seconded to Base4 to assist with development.
 
Title Pre-fertilisation diagnostics 
Description Method allows chromatid-level analysis of aneuploidy in human oocytes prior to fertilisation. 
Type Diagnostic Tool - Non-Imaging
Current Stage Of Development Early clinical assessment
Year Development Stage Completed 2009
Development Status Under active development/distribution
Impact The method continues to be evaluated by retrospective analysis of human subjects undergoing fertility treatment. Update Nov2012: the method is now applied to _prospectively_ to patients undergoing IVF at Europe's largest fertility centre. Early results are successful. I am developing software for the automated interpretation of results. 
 
Title Sequencing technology 
Description (See corresponding entry under IP) 
Type Diagnostic Tool - Non-Imaging
Current Stage Of Development Initial development
Year Development Stage Completed 2012
Development Status Under active development/distribution
Impact See above, and corresponding entry under IP. I would also like to point out that ResearchFish seems biased toward healthcare/point of care applications, and doesn't always fit with other research. 
 
Description Teaching/mentoring/advising - Malaysia 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Type Of Presentation Workshop Facilitator
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact I teach and mentor regularly at the Universiti Kebangsaan, Malaysia. I teach a workshop in scientific paper writing, and also a workshop in science strategy and innovation (both to postgraduates, and postdocs). I also periodically advise senior officials of the University and (less often) Malaysia's science ministry. This takes place annually.

See above.
Year(s) Of Engagement Activity 2008,2009,2010,2011,2012