Dr Joseph Hoolachan

Entry level post-doctoral researcher for the skeletal muscle team of Dr Debbie Thurmond lab in the Arthur Riggs Diabetes & Metabolism Research Institute at City of Hope. Focused on prevention/reversal of skeletal muscle insulin resistance and mitochondrial dysfunction in diabetes. Projects include investigating novel protein targets for reversal/prevention of mitochondrial damage, cross collaboration of investigating TAS1R3 as a multitasker target in pancreatic beta cells and skeletal muscle, cross collaboration of novel small molecule inhibitory drug for diabetes. Culture of L6.GLUT4myc (murine) and LHCN-M2 (human) immortalized myoblast and myotubes. Cell culture tools: Pharmacological treatment, siRNA transfection for gene knock down, plasmid transfection for gene overexpression in myoblasts, MTT assay for cell viability, LDH assay for cytotoxicity, Tritium labelled 2-DG uptake assays, GLUT4myc immunostaining assays in L6.GLUT4myc myoblasts, insulin stimulation and palmitate treatment to emulate lipotoxicity. In vitro: RNA isolation (commercial and Trizol-chloroform), protein isolation (NP40 lysis), reverse transcription, DNA isolation, qPCR, western blot. Animal handling: Colony management, genotyping, doxycycline treatment for inducible transgenic mice, mouse dissection, gavage, intraperitoneal and intramuscular injections. Phenotypic testing of weight, glucose and insulin tolerance test. Writing and data analysis: Capability for writing scientific papers, reviews and grant. Critical reviewing for manuscripts. Experienced in developing presentations for various levels of meetings and conferences. Versed in statistical analysis for GraphPad PRISM and intermediate for statistical analysis in R. Personal Skills: Organization, time management, independent work ability alongside team work for collaborative projects. Learning and developing skills for implementing single nuclear RNA-Sequencing to address biological questions in mature muscle fibers.

• Processing of clinical samples in Class 2 Biological Safety Cabinet. • Viral RNA extraction and purification by magnetic particle separation using KingFisherTM Flex Purification System (ThermoFisher Scientific) • Maintaining a sanitary lab environment within a UKAS ISO17025 accredited library. • Ensuring efficient turnaround of sample processing for quick results to clients. • Teamwork and communication to ensure optimal sample processing.

Prerequisites 1. Completed necessary pre-requisite of Introduction to Teaching and Demonstrating (ITAD) course at Keele University. Role and Responsibilities 1. Assisted the main demonstrator(s) in the supervision of first- and second-year medical students in laboratory sessions that covered different disciplines such as physiology, biochemistry, histology, microbiology and genetics. 2. Interacted with students in professional and pastoral manner to provide support with any difficulties, discuss the experiment and aims and promote their free thinking in tackling the aims and objectives. 3. Assessed student’s knowledge and understanding of the experiments and subject content through open ended questions to promote independent learning. 4. Monitored for any health and safety risks in each session. Awards and/or Continuing Professional Development 1. Awarded internal qualification of ITAD at Keele University 2. Awarded Associate Fellowship with Advance HE.

Role Involved in the logistical processing of pre-examined and examined AQA GCSE and A level exam papers. Responsibilities 1. Allocated exam papers to specific examiners and confirmed exam paper retrieval through company internal database. 2. Implemented IT skills to process details of examiners, students and marked final grades into the AQA internal EPS database. 3. Used time management and organisational skills to reach daily quota of >1000 exam papers with complete accuracy to ensure delivery to correct examiner and no overbookings. 4. Engaged in teamwork and communication to ensure any minimisation of logistical error.

Role I worked as a temporary project assistant for an external company that aimed to improve the office efficiency for their client. Responsibilities 1. Data collection of office environment by observational analysis. 2. Use of Microsoft Excel for data input. 3. Time management and organisation to ensure data collection occurred at hourly intervals. 4. Engaged in communication with office staff for dealing with queries regarding the purpose of the study. 5. Collected raw data through observational analysis in an office setting to aid in a study for improving office efficiency. 6. Developed experience of independent working.

This was a week long summer school spent in the Hawkhills, York run by the Gatsby Foundation.. During this week a series of lectures and workshops on plant science was ran; the aim of this was to interest first year biological sciences students into plant science. Tasks • Attended lectures on current areas of plant science research which included: plant ecosystem modelling, plant developmental biology, stomata research. optigenetic approaches on plants, using plants for drug development and protecting plants from pathological diseases. • Participated in Q&A seminars of the lectures. • Use computer modelling to understand plant development. • Use gene sequence analyser to determine the pathological disease infecting a certain plant. • Develop taxonomic skills for categorising and finding species of plants. • Use a confocal fluorescence microscope and subsequent computer software to visualise images of a leaf cell of Arabidopsis thaliana. • Carry out an ecological experiment regarding effect of leaf temperature on size.

Thesis: A transcriptomic-based drug repositioning approach for the identification of novel specific therapies for spinal muscular atrophy. https://eprints.keele.ac.uk/id/eprint/10744/ Bioinformatics: Proficiency in QC analysis, alignment and differential gene expression analysis for transcriptomics in Galaxy. Intermediate in R coding. Versed in pathway analysis and biological/drug databases. Lab skills: Murine (C2C12) and human (LHCN-M2) immortalised muscle cell culture, RNA isolation (commercial and Trizol-chloroform), reverse transcription, qPCR, PCR, siRNA transfection, agarose gel electrophoresis, in vitro drug treatment, LDH assay (cell toxicity), BrDU labelling assay (cell proliferation). Animal Handling: General husbandry, weaning, genotyping and colony management of wild type and transgenic mice. Drug administration via gavage and intraperitoneal routes, phenotypic measurement of weight and motor function (righting reflex) and dissection for tissue harvesting.

Modules Advanced Statistics, Research Methods, Introduction to Research, Advanced Molecular Medicine, Bioinformatics, Post-Genomic Bioinformatics, Professional Skills, MSc Research Project Lab Techniques Mammalian Cell Culturing Cancer Cell Culturing MTT based Cell Viability Assay Agarose Gel Electrophoresis PCR Restriction Digest In Vitro Transcription DNA Extraction Computer Skills R coding software for univariate and multivariate statistical analysis. Docker coding software for bioinformatics analysis. Biolinux operating system for conducting bioinformatics analysis. Sequence Alignment: NCBI Blast, DIALIGN, MUSCLE, CLUSTAL, Jalview Genomics: Artemis, ACT Protein Structure: PyMol, ProSA, VERIFY_3D Databases: KEGG, UniProt, NCBI, HMDB Gene Expression: DeApp, EdgeR, DAVID 6.8, WebGestalt Project Investigation of Duplex Sequencing for ex vivo comparison of DNA damage in the RAS isoforms.

Modules Core: Classical Genetics, Molecular Genetics, Genetic & Genomic Technologies, Biochemistry, Molecular Cell Biology Application of Genetics: Medicine, Plant Science, Microbiology, Evolutionary Biology Non-Scientific Subjects: Bioethics, History of Science, Leadership, Entrepreneurial Skills Lab Techniques Developed Amoeboid Culturing RNA extraction cDNA conversion Agarose Gel Electrophoresis PCR RT-qPCR Computer Skills Statistical Analysis: GraphPad PRISM Sequence Alignment: NCBI Blast Primer Design: GeneScreen Viewer Project Investigated the genotype-phenotype relationship between growth rate of different Dictyostelium discoideum isolates on a diet of Klebsiella aerogenes. Used qPCR to measure gene expression of candidate genes involved in advantaged or disadvantaged growth in knock out D. discoideum mutants. Correlated gene expression data to phenotypic growth rates of the natural isolates when grown on media of Klebsiella aerogenes.