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Chloroplast genomes in the genus Passiflora: cp-DNA sequencing, annotation and comparative studies

Grant number: 19/09210-4
Support type:Scholarships abroad - Research Internship - Doctorate (Direct)
Effective date (Start): June 17, 2019
Effective date (End): December 16, 2019
Field of knowledge:Biological Sciences - Genetics
Principal Investigator:Maria Lúcia Carneiro Vieira
Grantee:Luiz Augusto Cauz dos Santos
Supervisor abroad: Helene Berges
Home Institution: Escola Superior de Agricultura Luiz de Queiroz (ESALQ). Universidade de São Paulo (USP). Piracicaba , SP, Brazil
Local de pesquisa : Institut National de la Recherche Agronomique, Toulouse (INRA), France  
Associated to the scholarship:17/04216-9 - Chloroplast genomes of Passiflora: structure, evolutionary inferences and gene transfer to the nucleus, BP.DD

Abstract

The family Passifloraceae consists of about 700 species of herbaceous or woody vines, shrubs and trees, classified in around 16 genera, with almost all of its members belonging to the genus Passiflora, popularly known as passion fruits or passionflowers. The Passiflora genus has been studied using morphological and molecular descriptors, and despite the recent taxonomical classification into four subgenera (Astrophea, Decaloba, Deidamioides and Passiflora), there are unsolved phylogenetic relationships in this group, mainly on the existence of the subgenus Deidamioides. However, the lack of DNA data is one of the reasons for the difficult to make phylogenetic inferences as well as carrying out genomic studies. With the objective to describe and understand the genome structure of passionflowers, our group has constructed a large-insert genomic library of P. edulis in collaboration with INRA at the French Plant Genomic Resources Center (CNRGV), and the exploitation of this library generated, for instance, the chloroplast (cp) genome sequence of P. edulis, the first chloroplast genome sequenced in the Passifloraceae family. Next, we proposed to analyze the cp-genome of several Passiflora species by chloroplast isolation and cp-DNA sequencing using the Single Molecule Real-Time (SMRT) technology, implemented by Pacific Biosciences (PacBio). However, to access the information on cp-genomes is every critical. The most critical step is the chloroplast DNA isolation without nuclear DNA contamination that usually adheres to the outer chloroplast membrane. In collaboration with CNRGV, in a previous 3-month internship (BEPE), we had successfully isolated and sequenced the chloroplast genomes of three Passiflora species: P. adenopoda (Decaloba), P. racemosa (Passiflora) and P. sprucei (Passiflora). Due to the necessity to continuing our research with more and diversified plant material, after coming back to Brazil, we have collected several species, and now we propose to restart the chloroplast isolation and sequencing the cp genomes using PacBio technology in collaboration with CNRGV. In addition, we will also optimize in France a CRISPR/Cas9 assay to capture cp-DNA fragments from Passiflora species, a very innovative tool to be applied in chloroplast genome studies. Finally, the data generated through this BEPE will lend support to the proposed doctoral plan, that is, to investigating different aspects of the Passiflora chloroplast genome structure and evolution.