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Identification of passion fruit (Passiflora edulis) chromosomes using BAC-FISH

  • M. A. Sader
  • Y. Dias
  • Z. P. Costa
  • C. Munhoz
  • H. Penha
  • H. Bergès
  • M. L. C. Vieira
  • Andrea Pedrosa-HarandEmail author
Original Article

Abstract

Passiflora edulis, the yellow passion fruit, is the main crop from the Passiflora genus, which comprises 525 species with its diversity center in South America. Genetic maps and a BAC (bacterial artificial chromosome) genomic library are available, but the nine chromosome pairs of similar size and morphology (2n = 18) hamper chromosome identification, leading to different proposed karyotypes. Thus, the aim of this study was to establish chromosome-specific markers for the yellow passion fruit using single-copy and repetitive sequences as probes in fluorescent in situ hybridizations (FISH) to allow chromosome identification and future integration with whole genome data. Thirty-six BAC clones harboring genes and three retrotransposons (Ty1-copy, Ty3-gypsy, and LINE) were selected. Twelve BACs exhibited a dispersed pattern similar to that revealed by retroelements, and one exhibited subtelomeric distribution. Twelve clones showed unique signals in terminal or subterminal regions of the chromosomes, allowing their genes to be anchored to six chromosome pairs that can be identified with single-copy markers. The markers developed herein will provide an important tool for genomic and evolutionary studies in the Passiflora genus.

Keywords

Bacterial artificial chromosome (BAC) Fluorescent in situ hybridization (FISH) Karyotype Cytogenetic map Repetitive DNA sequences Retrotransposon 

Abbreviations

BACs

Bacterial artificial chromosomes

FISH

Fluorescent in situ hybridizations

NORs

Active nucleolar organizer regions

CMA

Chromomycin A3

DAPI

4′,6-Diamidino-2-phenlindol

rDNA

Ribosomal DNA

TEs

Transposable elements

SSRs

Microsatellite sequences

PCR

Polymerase chain reaction

SSC

Saline–sodium citrate buffer

SDS

Sodium dodecyl sulfate

Cy3-dUTP

5-Amino-propargyl-2′-deoxyuridine 5′-triphosphate coupled to red cyanine fluorescent dye

Alexa

Alexa Fluor 488

FITC

Fluorescein isothiocyanate

BSA

Bovine serum albumin

LINE

Long interspersed nuclear element

Notes

Acknowledgments

We thank Mr. Steve Simmons for proofreading the manuscript.

Author contribution

MAS and APH conceived and designed the research. HB, HP, and MLCV contributed to BAC library, plant material, and bioinformatic data. MAS and YD conducted the experiments. CM, MAS, YD, and ZPC analyzed the data. MAS and APH wrote the manuscript. All authors read and approved the manuscript.

Funding information

The authors received financial support from CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico). Scholarships from CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) were awarded to MA Sader, ZP Costa, and A. Pedrosa-Harand. This study was supported in part by the Coordenacão de Aperfeicoamento de Pessoal de Nível Superior - Brasil (CAPES, Finance Code 001), and Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP grant no. 2014/25215-2).

Supplementary material

10577_2019_9614_Fig4_ESM.png (8.6 mb)
Figure S1

Fluorescent in situ hybridization to P. edulis chromosomes. (a) BAC Pe134H15 (red); (b) Pe164A12 (red) and 35S rDNA (green); (c) BAC Pe173B16 (green) and Pe164M13 (red); (d) BAC Pe195F4 (red) and 35S rDNA (green); (e) BAC Pe173B16 (red) and 35S rDNA (green). Chromosomes were counterstained with DAPI, are visualized in grey and indicated by number. BACs were labeled with Cy3-dUTP and pseudocolored. Bar = 5 μm. (PNG 8848 kb)

10577_2019_9614_MOESM1_ESM.tif (19.9 mb)
High resolution image (TIF 20376 kb)
10577_2019_9614_MOESM2_ESM.docx (22 kb)
ESM 1 (DOCX 22 kb)

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Department of BotanyFederal University of PernambucoRecifeBrazil
  2. 2.Department of Genetics, “Luiz de Queiroz” College of AgricultureUniversity of São PauloPiracicabaBrazil
  3. 3.Department of Technology, Faculty of Agricultural and Veterinary SciencesSão Paulo State University (UNESP)JaboticabalBrazil
  4. 4.French Plant Genomic Resources Center (CNRGV)/ INRAToulouseFrance
  5. 5.Laboratório de Citogenética e Evolução Vegetal, Departamento de Botânica, Centro de BiociênciasUniversidade Federal de PernambucoRecifeBrazil

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