The Functional Genomics Core Facility (FGCF)
The Functional Genomics Core Facility (FGCF), directed by Prof. Roberto Piva, was established at the MBC Torino in 2012 as a result of a collaborative project involving three research centres (CeRMS, MBC, HuGEF), thanks to the contributions of Fondazione di San Paolo and Italian Ministry of Health.MissionThe main goal of the FGCF is to facilitate the application of high-throughput functional screening approaches to biomedical research by providing a platform of up-to-date instrumentations and adequate expertise to the research community. The FGCF participates in several collaborative research projects aimed to enlighten biological processes that fall into the broad concept of Loss Of Function (LOF) and Gain Of Function (GOF) screening technologies.The FGCF is in charge of designing and coordinating the experiments carried out in the Functional Genomics Core Facility, as well as with immunocompromised (NSG) mice. Progresses are constantly monitored by monthly reports and meetings with Collaborators aimed at the advancement of each specific task and the improvement of the research plan. An external referee monitors FGCF activities and provide external strategic advice.Functional screening approaches are currently used to identify putative biomarkers of clinical drug response/resistance and to guide the therapeutic decision-making process in several lymphoma models. Screening are executed using medium-scale lentiviral shRNA libraries designed to target key regulators of pathological cancer homeostasis including genes involved in cell proliferation, survival, metabolism, DNA damage protection, migration, invasion, inflammation, and other core cancer pathways.In order to recognize genes actively sustaining insensitivity to specific inhibitors (i.e. ALK, mTOR, PI3K, proteasome), the biological effects of each gene knockdown is monitored on multiple assays such as proliferation, survival/apoptosis, morphology, migration, invasion, and in vivo tumor growth in the presence or absence of the different drugs.
The FGCF hosts automated platforms (2 STARlet workstations, Hamilton) to screen genetic (shRNA, siRNA, CRISPRgRNA, cDNA) and chemical libraries, perform high-throughput quantitative realtime PCR (CFX 384, Biorad), and analyse phenotype read-outs through a multi-mode plate reader (Synergy 2, Biotek). NSG mice required for these studies are housed at the MBC mouse facility.
TRC shRNA sub-library (Cancer Genes v1.0)
- Human Kinase Open Reading Frame Collection (559 human kinases ORFs from Broad Institute-MIT)
- Genome-wide pooled lentiviral human shRNA library (~200,000 shRNA constructs from SBI)
- Mouse testis cDNA pooled retroviral library (Stratagene)
- Lentiviral Genome-scale CRISPR-Cas9 KnockOut (GeCKO) library (64,751 gRNAs targeting 18,080 genes from Broad Institute-MIT)
- Perform low-throuput to genome-wide RNAi, microRNA, cDNA, gRNA and chemical screenings in human/mouse cells
- Production of lentiviral paricles validated for On-target and Off-target effects by RT-qPCR and/or gene expression profiling
- Generation of stable cell line expressing constitutive or inducible shRNA/cDNA
- Production of custom sublibraries
- Nucleic acid extraction (genomic DNA, plasmid DNA, RNA)
- RT-qPCR
The FGCF welcomes collaboration to perform target and genomewide screenings using high-throughput technologies. Research groups external to MBC/ CeRMS are welcome to contact the FGCF concerning collaborative projects and training opportunities.Although priority is given to screening projects, the FGCF also collaborates in small-scale research projects that benefit from the use of the available equipment and accumulated expertise in automation, assay development and data acquisition/analysis.
- Roberto Piva- Associate Professor
- Garaffo Giulia - Postdoctoral Fellow
- Pellegrino Elisa- Postdoctoral Fellow
- Nicoletta Vitale - Postdoctoral Fellow
- Elisabetta Mereu - Postdoctoral Fellow
- Scarfo' Irene - Postdoctoral Fellow
- Maria Moscvin - MD/PhD Student
- Elisa Bergaggio - Fellow
- Cecilia Bandini - Fellow
- Carlotta Duval - Graduate Student
- Elisa Setten - Graduate Student