Potential of application of post edited cells modified TFAM gene by CRISPR Cas 9 technology in bovine model

• Grant number: 17/08896-4
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: December 01, 2017
• End date: June 30, 2021
• Field of knowledge: Agronomical Sciences - Veterinary Medicine - Animal Reproduction
• Principal Investigator: Carlos Eduardo Ambrósio
• Grantee: Vanessa Cristina Oliveira Nogueira de Pontes
• Host Institution: Faculdade de Zootecnia e Engenharia de Alimentos (FZEA). Universidade de São Paulo (USP). Pirassununga , SP, Brazil
• Associated scholarship(s): 19/04442-4 - Knock-in of human TFAM gene in bovine fibroblasts cells by CRISPR/Cas9 technology, BE.EP.PD

Abstract

The mitochondrial transcription factor A (TFAM) is a member of HMGB subfamily that binds to promoters of mtDNA. It is a very important gene that maintains mtDNA, regulates the number of copies and is essential for the initiation of transcription mtDNA. Recently, gene edition techniques have been used as a very effective tool in genomic manipulation. The new technology called CRISPR/Cas9 (Regulary interspaced clustered short palindromic repeats) uses a short gRNA containing 20 nucleotides complementary to the DNA sequence. When gRNA binds to the target site, the Cas9 protein is recruited to bind in the chosen location and induce double strands breaks in DNA. In this context, we precisely edited the TFAM gene in bovine fibroblasts by CRISPR Cas 9 technology, once this gene regulates the replication specificity of the mtDNA, consequently aiming to generate a Rho zero cell. Thus, this proposal aims to analyze if the post-edited cells will remain in culture, presents a decrease in the number of mitochondrial copies and a depletion of mtDNA of the potential membrane. The characterization expands a field of little exploration, allowing confirmation of the generation of a new study model of mitochondrial inheritance and segregation. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

DE OLIVEIRA, VANESSA CRISTINA; MARIANO JUNIOR, CLESIO GOMES; BELIZARIO, JOSE ERNESTO; KRIEGER, JOSE EDUARDO; BRESSAN, FABIANA FERNANDES; SANTOS ROBALLO, KELLY CRISTINE; FANTINATO-NETO, PAULO; MEIRELLES, FLAVIO VIEIRA; CHIARATTI, MARCOS ROBERTO; CONCORDET, JEAN-PAUL; et al. Characterization of post-edited cells modified in the TFAM gene by CRISPR/Cas9 technology in the bovine model. PLoS One, v. 15, n. 7, JUL 10 2020. (17/08896-4)

DE OLIVEIRA, VANESSA CRISTINA; ALVES MOREIRA, GABRIEL SASSARAO; BRESSAN, FABIANA FERNANDES; MARIANO JUNIOR, CLESIO GOMES; SANTOS ROBALLO, KELLY CRISTINE; CHARPENTIER, MARINE; CONCORDET, JEAN-PAUL; MEIRELLES, FLAVIO VIEIRA; AMBROSIO, CARLOS EDUARDO. Edition of TFAM gene by CRISPR/Cas9 technology in bovine model. PLoS One, v. 14, n. 3, MAR 7 2019. (17/08896-4, 13/08135-2)