GT training- A GOSH experience

Sumera Ghani

Great Ormond St Hospital London

Paediatric Malignancy Unit

About Me

Sept. 2003- July 2006- BSc in Human Genetics University College London (UCL)

Sep. 2007- Oct. 2008- MSc in Experimental Oral Pathology at Queen Mary University of London

August 2008- Started working as Genetic Technologist at Great Ormond St Hospital NHS trust

About the lab CPA accredited lab

UK NEQAS

The team comprises of a: Consultant Cytogeneticist Section Head/Principal Molecular Geneticist Principal Clinical Cytogeneticist Chief Biomedical Scientist Senior Clinical Cytogeneticist 3 Clinical Cytogeneticist 2 Medical Technical Officers

PMU OVERVIEW.

PMCU- Diagnostic Cytogenetics Unit

We work closely with the haematology/oncology department and their consultants within GOSH.

PMCF- Genetics Core Facility New facility established in September 2008 Provide both Molecular Genetics and Cytogenetics tests to: NHS Trusts Academic Institutions Private Labs Pharmaceutical Industries

PMCU- Cytogenetic training Booking in samples- Solid tumour Blood Bone Marrow

Lymph nodes Setting up cultures Harvesting Slide making Banding

Techniques Learnt: G banding Culturing Solid tumours and preparing slides. Setting up FISH and analysis Auditing Extraction of Mononuclear cells for freezing in liquid Nitrogen DNA and RNA extraction Agarose Gel analysis Setting up a PCR reaction Sequencing Growing Fosmids and BACS Extracting DNA from Fosmids and BACS Nick translation- labelling DNA

FISH (Fluorescent In Situ Hybridisation)

SETTING UP FISH Paraffin Embedded Slides Touch Imprints Bone Marrow Solid tumour Blood

DIFFERENT TYPES Whole Chromosome Paints LSI FISH Metaphase FISH

FISH (Fluorescent In Situ Hybridisation)Locus specific probe

t(9;22)(q34;q11).

In addition to chromosome analysis (karyotyping)

a full range of adjunctive tests, such as Fluorescence in Situ Hybridisation (FISH),

Reverse Transcriptase polymerase chain reaction (RT-PCR) for translocation detection of specific fusion genes.

Mutation detection for a number of genes using direct sequencing, genotyping and MLPA.

PMCF- Genetics core facility

PMCF- Genetics core facility DNA and RNA extraction from: Blood Bone marrow Fixed material Solid tumour Paraffin embedded material

DNA Integrity/concentration

RNA AND DNA QUALITYLADDER

DNA1

DNA2

DNA3

RNA1

RNA2

RNA3

PCR

Overall experience

Core skills gained in both Molecular and Cytogenetics

Transferable skills

Opportunity to rotate and continue to develop

Opportunity to continue personal learning

Broad experience

Lab project

Questions?

Methods of extracting DNA/RNA

Flexigene- DNA Trizol/chloroform-RNA Phenol/chloroform-DNA Ambion-DNA

PCR

Samples that are accepted for cytogenetic investigations include:

ALL  AML JMML/CML  MDS  Other leukaemia TAM Congenital haematological conditions Infiltrating tumours Post-treatment remission samples will be accepted if a prior cytogenetic abnormality

has been detected. Additionally, pleural fluid or CSF diagnostic and relapse specimens may be accepted.

ELA2 NEUTROPHILS