My project is entitlted "Molecular regulation of transcription termination". In this project we routinely use CRISPR to tag porteins of interest (CPSF30, POLR2A), generating stable cell lines. We use a variety of tagged cell lines (GFP, mAID, mTURBO) in order to study the mechanism of transcriptional termination. For example we couple auxin inducible degradation (AID) of specific proteins (CPSF30/73 etc) with RNAseq to establish what occurs to transcription when CPSF30/73 is acutely depleted.

Shortly after red blood cells (RBCs) develop from reticulocytes into mature erythrocytes, all ribosomal RNA, messenger (m)RNA and transfer (t)RNA are destroyed by programmed ribonucleolysis, with no further protein production occurring in these cells. As miRNA regulate translation, it is therefore surprising to find a large pool, of stable, catalytically active miRISC within mature RBCs. As there is no canonical function for miRNA in RBCs, I investigated the possibility that they functioned in a novel manner, or, they functioned in macrophages following the routine phagocytosis that occurs at the end of an RBCs lifespan. During my PhD I: * Investigated what interacts with RBC miRISC via antibody or 2’O-methyl based pull down * Generated monocyte derived macrophages from healthy donors (blood transfusion service buffy coats)* Generated a method of characterising RBC microvesicles by flow cytometry * Established an RBC / RBC-microvesicle phagocytosis model * Performed RNAseq in primary macrophages to establish how they respond, at a transcrip level, to phagocytosing RBC microvesicles * Investigated erythrocytic miRNA function in macrophages via luciferase assays

Interferons were originally perceived to be important mediators of the immune response, acting on mature cells. They were also shown to have anti-proliferative properties and were therefore used in the treatment of myeloproliferative disorders. New data indicates a role for IFNs in haemopoietic homeostasis, with IFNs having a stimulatory effect on quiescent hematopoietic stem cells. During my project with Dr Helen Wheadon I demonstrated that IFN-α stimulation appeared to cause hESC to proliferate while IFN-γ stimulation caused them to differentiate towards an endothelial lineage. * Evaluate whether ES cells respond to IFNs. * Determine the effect of IFN treatment on morphology, differentiation and proliferation of ES cells. * Profile myeloid lineage priming following IFN treatment of early haematopoietic progenitors.

Masters in Molecular Medicine at the University of Glasgow where I had two projects:"Expression profiling of CML CD34+ stem cells"* Expression profiling of CML patient samples to identify potential targets* Verifying expression data via immunoblotting * Evaluating the response of identified targets to current therapies in vitro "Evaluating the distribution of RNA polymerase II transcripts"* Observed the distribution of nascent RNA in vitro by labelling incorporated 5-ethynyl uridine with azide modified dyes via Click Chemistry* Developed a method to isolate poly(A) RNA from cytoplasmic and nucleoplasmic RNA pools * Prepared biotinylated cDNA probes from isolated nuclear and cytoplasmic poly(A) RNA * Observed the distribution of poly(A) RNA and rRNA in mammalian cells using oligo dT and cDNA FISH

Developing an in vitro diagnostic device (IVD) that coupled molecular biology with surface enhanced resonance Raman scattering (SERRS) to detect viral, bacterial and fungal pathogens. During my time at Renishaw I: * Optimised the multiplex PCR portion of the Renishaw IVD * Designed and executed studies to improve the specificity and sensitivity of the Renishaw IVD * Created GMP grade plasmid and glycerol stocks for each target detected by the Renishaw IVD * Designed and managed a sequencing study with MWG Operon

Developing electrophoretic and immunoblot assays for qualitative analysis of the Japanese encephalitis virus (JEV) vaccine Ixiaro™. During my time at Intercell I: * Developed a slot blot assay to semi-quantitate the residual amount of vero cell protein in manufactured vaccine (used as a QC test before sale)* Designed and executed study protocols to resolve the needs of the manufacturing department