Induction of mutations in the Aire gene by means of CRISPR-Cas9 system in mTEC 3.10 cells

• Grant number: 20/01879-0
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: April 01, 2020
• End date: November 30, 2021
• Field of knowledge: Biological Sciences - Genetics - Molecular Genetics and Genetics of Microorganisms
• Principal Investigator: Geraldo Aleixo da Silva Passos Júnior
• Grantee: Letícia Agrelli de Brito
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated research grant: 17/10780-4 - Effect of CRISPR-Cas9-induced mutations in the Aire gene (APS1 syndrome) on protein conformation, mTEC cell transcriptome and their interaction with thymocytes, AP.TEM

Abstract

The IC fellow will work with the graduate and post-doctoral students of the thematic project under supervision. She will use the CRISPR-Cas9 system to induce mutations in the Aire gene, in mTEC cells line 3.10, the same mutations that are associated with the APS1 syndrome. We will make the selection of the mutations of the human Aire gene (SAND and PHD domains) to design the guide RNA (gRNA) of the CRISPR-Cas9 system, following the transfections of mTEC cells in culture and selection and characterization of the Aire mutant clones, separation of single cells by means of FACS and molecular identification of mutant clones by Sanger sequencing.