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Use of CRISPR / Cas9 in the editing of susceptibility genes of Citrus sinensis to Xanthomonas citri subsp. Citri

Grant number: 17/13885-1
Support Opportunities:Scholarships in Brazil - Doctorate
Start date: December 01, 2017
End date: December 01, 2021
Field of knowledge:Agronomical Sciences - Agronomy
Principal Investigator:Alessandra Alves de Souza
Grantee:Reinaldo Rodrigues de Souza Neto
Host Institution: Instituto Agronômico (IAC). Agência Paulista de Tecnologia dos Agronegócios (APTA). Campinas , SP, Brazil
Associated research grant:13/10957-0 - Xylella fastidiosa-vector-host plant interaction and approaches for Citrus Variegated Chlorosis and citrus canker control., AP.TEM
Associated scholarship(s):18/21990-2 - Use of CRISPR-Cas9 to edit citrus canker susceptibility genes to obtain commercial sweet orange varieties resistant to Xanthomonas citri subsp. citri, BE.EP.DR

Abstract

Citrus canker is a disease caused by the bacterium Xanthomonas citri subsp. citri (X. citri) and it has been associated to important economic losses in citrus orchards at worldwide. The symptoms of citrus canker are characterized by eruptive lesions that form on the surface of leaves, branches and fruits. With the development of the disease, there are the abscission of leaves end premature fall of fruits, reducing the plant production. It is known that the pathogenicity of X. citri is depend on translocation of effector proteins known as Transcription Activator-Like Effectors (TALEs) to the inside of plant cell. TALEs working as transcription factor in host cell and binding to Effector Binding Elements (EBE) on promoters of specific plant genes activating its expression. These genes are denominated of "susceptibility genes" because their expression promote to development of disease and bacterial colonization. PthA4 is a TAL effector of X. citri essential to induction of hypertrophy and hyperplasia in citrus. It is known that PthA4 trans-activate LOB1 gene in sweet orange, whose expression is correlated with citrus canker development. However, the only expression of LOB1 is not enough to generate citrus canker symptoms, suggesting that other activated genes by PthA4 will be necessary to development of disease. Thus, such genes became good targets to genome edition aiming to obtain resistant plants to X. citri. Therefore, the goal of this study is identifying new susceptibility genes to X. citri aiming to edit their EBE regions using the CRISPR/Cas9 system. Based on previously transcription data and bioinformatic analysis, eight genes of sweet orange were selected, which are showing the target sequence of PthA4 in their promoter regions. The relative expression of these genes will be evaluated in different varieties of sweet orange. The most induced genes, whose expression be depends on PthA4, will be selected to edition by CRISPR/Cas9 by transient expression and genetic transformation. This project intends to identify new direct targets of PthA4in commercial varieties of sweet orange and obtaining edited resistant plants by CRISPR/Cas9 to X. citri.

News published in Agência FAPESP Newsletter about the scholarship:
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VEICULO: TITULO (DATA)
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Scientific publications (4)
(The scientific publications listed on this page originate from the Web of Science or SciELO databases. Their authors have cited FAPESP grant or fellowship project numbers awarded to Principal Investigators or Fellowship Recipients, whether or not they are among the authors. This information is collected automatically and retrieved directly from those bibliometric databases.)
DE LIMA, LUIZ FELIPE FRANCO; CARVALHO, ISIS GABRIELA BARBOSA; DE SOUZA-NETO, REINALDO RODRIGUES; DOS SANTOS, LARISSA DA SILVA; NASCIMENTO, CESAR AUGUSTO; TAKITA, MARCO AURELIO; TAVORA, FABIANO TOUZDJIAN PINHEIRO KOHLRAUSCH; MEHTA, ANGELA; DE SOUZA, ALESSANDRA ALVES. Antisense Oligonucleotide as a New Technology Application for CsLOB1 Gene Silencing Aiming at Citrus Canker Resistance. PHYTOPATHOLOGY, v. 114, n. 8, p. 8-pg., . (21/09689-8, 17/13885-1, 20/04795-1, 19/23217-1, 22/04112-7)
DE SOUZA-NETO, REINALDO RODRIGUES; CAVALCANTE, LIDIA NASCIMENTO; CARVALHO, ISIS GABRIELA BARBOSA; CURTOLO, MAIARA; BENEDETTI, CELSO EDUARDO; TAKITA, MARCO AURELIO; WANG, NIAN; DE SOUZA, ALESSANDRA ALVES. CRISPR/Cas9-Mediated Disruption of CsLIEXP1 Reveals Expansin as a Key Susceptibility Factor for Citrus Canker Disease. MOLECULAR PLANT-MICROBE INTERACTIONS, v. 38, n. 4, p. 10-pg., . (23/05118-1, 17/13885-1, 13/10957-0, 14/50880-0, 20/04795-1, 18/21990-2)
DE SOUZA-NETO, REINALDO RODRIGUES; BARBOSA CARVALHO, ISIS GABRIELA; MOREIRA MARTINS, PAULA MARIA; PICCHI, SIMONE CRISTINA; TOMAZ, JUAREZ PIRES; CASERTA, RAQUEL; TAKITA, MARCO AURELIO; DE SOUZA, ALESSANDRA ALVES. qsR toxin as a biotechnological tool for plant pathogen bacterial contro. SCIENTIFIC REPORTS, v. 12, n. 1, . (17/13885-1, 17/16142-0, 13/10957-0, 16/15741-4, 14/50880-0, 16/01273-9)
DE SOUZA-NETO, REINALDO RODRIGUES; DA COSTA VASCONCELOS, FERNANDA NOGALES; TEPER, DORON; BARBOSA CARVALHO, ISIS GABRIELA; TAKITA, MARCO AURELIO; BENEDETTI, CELSO EDUARDO; WANG, NIAN; DE SOUZA, ALESSANDRA ALVES. The Expansin Gene CsLIEXP1 Is a Direct Target of CsLOB1 in Citrus. PHYTOPATHOLOGY, v. 113, n. 7, p. 12-pg., . (17/13885-1, 18/21990-2, 20/04795-1, 13/10957-0, 14/50880-0)
Academic Publications
(References retrieved automatically from State of São Paulo Research Institutions)
SOUZA NETO, Reinaldo Rodrigues de. Caracterização de CsLIEXP1 como um novo gene de susceptibilidade a Xanthomonas citri subsp. citri e edição gênica por CRISPR/Cas9 para tolerância ao cancro cítrico. 2022. Doctoral Thesis - Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia Campinas, SP.