Developing molecular biology kits for more sensitive diagnostics and gene expression analysis
Lead Research Organisation:
University of East Anglia
Department Name: Biological Sciences
Abstract
Molecular biology kits are widely used for research and diagnostics and are a multibillion dollar industry. Many of these kits use enzymes that modify DNA and RNA. Improving the accuracy and sensitivity of these enzymes can provide a kit with the unique selling point. In BBSRC-funded research, we have developed a novel approach to study DNA or RNA modifying enzymes using Next Generation Sequencing (NGS). This rapid, low-cost method was used to identify factors that improve the activity of DNA polymerases used in PCR, and RNA ligases used in kits that capture small RNAs (sRNAs). We plan to develop a High-Definition (HD) method that more accurately captures RNA molecules, which have been linked with cancers and disease. Funding is also requested to develop a PCRmax technology to generate diagnostic kits with increased sensitivity. PCRmax technology is a rare opportunity to develop a method leading to a step change in PCR efficiency.
Organisations
People |
ORCID iD |
| Tamas Dalmay (Principal Investigator) | |
| James Gray (Co-Investigator) |
Publications
Donaszi-Ivanov A
(2013)
Small RNA analysis in Sindbis virus infected human HEK293 cells.
in PloS one
Sorefan K
(2012)
Reducing ligation bias of small RNAs in libraries for next generation sequencing.
in Silence
Xu P
(2014)
Small RNA profile in moso bamboo root and leaf obtained by high definition adapters.
in PloS one
| Description | We discovered that ligation of small RNAs to adapter molecules is biased and that this bias is determined by the ability of a small RNA to anneal to the adapter molecule. With hindsight, it is not surprising because it is common sense that two molecules are more likely to be ligated to each other if they are physically next to each other. If a small RNA can anneal to an adapter, their ends are close to each other and have a better chance to get ligated than molecules that are not annealed to each other. Current commercial kits contain adapters with a fixed sequence therefore some sRNAs can anneal them and others can't, introducing a bias at the ligation step. We developed an approach that uses a pool of adapter sequences (by introducing degenerate nucleotides into the adapter) therefore more sequences can anneal to at least one of the adapter sequences. |
| Exploitation Route | We secured a European patent and also in the USA. Illumina decided not to licence the technology so we are negotiating with other companies. |
| Sectors | Agriculture, Food and Drink,Pharmaceuticals and Medical Biotechnology |
| Description | Our European Patent application has been granted in 2016 and in the autumn 2017 our US patent application was also approved. We have started negotiation with various companies about licencing. |
| First Year Of Impact | 2018 |
| Sector | Pharmaceuticals and Medical Biotechnology |
| Impact Types | Economic |
| Title | ANALYSING SEQUENCING BIAS |
| Description | The present invention relates to methods for determining the sequence bias of a sequencing technique. Furthermore, the invention relates to methods to reduce or enhance sequence bias during sequencing of nucleic acids via techniques involving adaptor ligations. Specifically the method relates to use of a degenerate RNA sequence to analyse sequence bias when generating small RNA libraries, and to the use of modified adaptors for cloning of small RNAs with degenerate or specific sequences to reduce or enhance sequencing bias, as well as various nucleic acid molecules relating thereto or derived therefrom. |
| IP Reference | WO2013017861 |
| Protection | Patent application published |
| Year Protection Granted | 2013 |
| Licensed | No |
| Impact | A company is already selling a kit that uses our technology and we are in discussion with them about licencing |