Genomics

The Genomics group provides comprehensive and integrated genomics analysis for the Unit Programmes and external collaborative projects.

The group carries out a range of genomic analyses, and is fully equipped to extract, purify, quantify and QC RNA and DNA for further investigation.  Currently the facility has the following sequencers and array technology systems:

1) ThermoFisher Scientific (formerly LifeTechnology) Ion Torrent PGM™

2) Illumina MiSeq Sequencer

3) Agilent Microarray technologies

4) ThermoFisher Scientific QuantStudio™ 6 Flex Real-Time PCR System (96- and 384-well capability)

The group provides support, training and analysis for a range of applications, including gene expression analysis, miRNA analysis, qPCR, ChiP-Seq, small genome and targeted genome sequencing (off-the-shelf and custom panels), polysome profiling and ribosome profiling.

Polysome profiling

Lysed cell or tissue samples are added to a sucrose gradient, ultracentrifuged and fractionated. This separates actively translated mRNAs from non-actively translated mRNAs. A UV trace provides the first indication that separation has been successful and helps in the identification of the sub-polysomal and polysomal regions.
RNA from individual fractions can be analysed using qPCR or pooled into "subs" and "polys" groups for microarray analysis.
Changes in the proportional representation of an mRNA across the gradient indicates a change in translational activity.

Bioinformatic support for microarrays is provided using GeneSpring software (Agilent technologies).

For more advanced informatics of sequencing data the Genomics group works closely with members of the Bioinformatics group.

Heat map

Following microarray analysis (gene expression or miRNA), data is analysed using GeneSpring and results are represented visually using hierarchial clustering, or heat maps.

In addition to our in-house sequencers, the Genomics group also prepare libraries (multiplexed if necessary) for ‘deep-sequencing’, through out-sourcing links with TGAC (The Genome Analysis Centre) in Norwich and the department of Biochemistry at the University of Cambridge. This provides access to ‘state-of-the-art’ Illumina HiSeq and NextSeq next generation Sequencers and allows any scale of project to be handled.

We have developed a pipeline for performing ribosome profiling.

Selected Publications

Design of nucleotide-mimetic and non-nucleotide inhibitors of the translation initiation factor eIF4E: Synthesis, structural and functional characterisation.

Soukarieh F, Nowicki MW, Bastide A, Pöyry T, Jones C, Dudek K, Patwardhan G, Meullenet F, Oldham NJ, Walkinshaw MD, Willis AE, Fischer PM.
Eur J Med Chem. 2016 Aug 24;124:200-217. doi: 10.1016/j.ejmech.2016.08.047.

Mitofusin-mediated ER stress triggers neurodegeneration in pink1/parkin models of Parkinson’s disease.

Celardo I, Costa AC, Lehmann S, Jones C, Wood N, Mencacci NE, Mallucci, GR, Loh SH, Martins LM.

Cell Death Dis. 2016 Jun 23;7(6):e2271. doi: 10.1038/cddis.2016.173.

mTORC1-mediated translational elongation limits intestinal tumour initiation and growth.

Faller WJ, Jackson TJ, Knight JR, Ridgway RA, Jamieson T, Karim SA, Jones C, Radulescu S, Huels DJ, Myant KB, Dudek KM, Casey HA, Scopelliti A, Cordero JB, Vidal M, Pende M, Ryazanov AG, Sonenberg N, Meyuhas O, Hall MN, Bushell M, Willis AE, Sansom OJ.

Nature. 2015 Jan 22;517(7535):497-500. doi: 10.1038/nature13896. Epub 2014 Nov 5.

Evaluating bias-reducing protocols for RNA sequencing library preparation.

Jackson TJ, Spriggs RV, Burgoyne NJ, Jones C, Willis AE.

BMC Genomics. 2014 Jul 7;15(1):569. doi: 10.1186/1471-2164-15-569.

Failure of translation of human adenovirus mRNA in murine cancer cells can be partially overcome by L4-100K expression in vitro and in vivo.

Young AM, Archibald KM, Tookman LA, Pool A, Dudek K, Jones C, Williams SL, Pirlo KJ, Willis AE, Lockley M, McNeish IA.

Mol Ther. 2012 Sep;20(9):1676-88. doi: 10.1038/mt.2012.116. Epub 2012 Jun 26.

 

Genomics

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