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, 10 (3), 447-463
eCollection

First Detailed Karyo-Morphological Analysis and Molecular Cytological Study of Leafy Cardoon and Globe Artichoke, Two Multi-Use Asteraceae Crops

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First Detailed Karyo-Morphological Analysis and Molecular Cytological Study of Leafy Cardoon and Globe Artichoke, Two Multi-Use Asteraceae Crops

Debora Giorgi et al. Comp Cytogenet.

Abstract

Traditionally globe artichoke and leafy cardoon have been cultivated for use as vegetables but these crops are now finding multiple new roles in applications ranging from paper production to cheese preparation and biofuel use, with interest in their functional food potential. So far, their chromosome complements have been poorly investigated and a well-defined karyotype was not available. In this paper, a detailed karyo-morphological analysis and molecular cytogenetic studies were conducted on globe artichoke (Cynara cardunculus Linnaeus, 1753 var. scolymus Fiori, 1904) and leafy cardoon (Cynara cardunculus Linneaus, 1753 var. altilis De Candolle, 1838). Fluorescent In Situ Hybridization In Suspension (FISHIS) was applied to nuclei suspensions as a fast method for screening of labelling probes, before metaphase spread hybridization. Classic Fluorescent In Situ Hybridization (FISH) on slide, using repetitive telomeric and ribosomal sequences and Simple Sequence Repeats (SSRs) oligonucleotide as probes, identified homologous chromosome relationships and allowed development of molecular karyotypes for both varieties. The close phylogenetic relationship between globe artichoke and cardoon was supported by the very similar karyotypes but clear chromosomal structural variation was detected. In the light of the recent release of the globe artichoke genome sequencing, these results are relevant for future anchoring of the pseudomolecule sequence assemblies to specific chromosomes. In addition, the DNA content of the two crops has been determined by flow cytometry and a fast method for standard FISH on slide and methodological improvements for nuclei isolation are described.

Keywords: Cynara; FISH; FISHIS; SSR simple sequence repeats; flow cytometry; repetitive sequences.

Figures

Figure 1.
Figure 1.
Flow Cytometry DNA content histogram. Flow cytometry analysis of DNA fluorescence peaks (FL1A) from G0/G1 (2C) PI stained leaf nuclei from cultivated cardoon (M1), globe artichoke (M2) and Pisum sativum (pea) (M3). Pea was used as an internal standard and the peak was set at channel 400; M4 shows pea G2 nuclei (4C).
Figure 2.
Figure 2.
The smallness and variable sizing of Cynara chromosomes is shown; in both panel A and B, leafy cardoon chromosome 1 has been circled as an example of the different condensation level of the same metaphase chromosomes. In A chromosome 1 is 3.2 µm, in B it is 4.3 µm (a 36% size variation). Scale bars: 5 µm.
Figure 3.
Figure 3.
Chromosome numbering. Metaphase spreads showing numbered chromosomes of cardoon (A) and globe artichoke (B) counterstained with DAPI. Scales bars: 5 µm.
Figure 4.
Figure 4.
FISH molecular cytogenetic analysis with rDNA. FISH on metaphase spreads of cultivated cardoon (A) and globe artichoke (B) using the rDNA probe pTa71-Cy3 (red fluorescence). In Fig. 3B, arrows point to the hybridization spots localized on the three small acrocentric chromosomes, placed head to head. Scale bar: 5 µm.
Figure 5.
Figure 5.
Fast screening of labelling probes was performed by FISHIS on cultivated cardoon (panel A and C) and globe artichoke (panel B and D) nuclei suspensions. The nuclei with the clearest and discrete telomeric (TTTAGGG)5 and SSR (GAA)7 signals are shown. All oligonucleotides were fluorescently labelled by Cy3 (red fluorescence). Nuclear DNA was counterstained with DAPI (blue fluorescence). Scale bar: 5 µm.
Figure 6.
Figure 6.
FISH molecular cytogenetic analysis with SSR probes. FISH on metaphase spreads of cardoon (left hand side) and of globe artichoke (right had side). The oligonucleotides sequence is indicated in each panel. Oligonucleotides were labelled with Cy3 (red fluorescence) or FITC (green fluorescence) fluorochromes. In (E) and (F) chromosomes 3, 5 and 8 of cardoon and globe artichoke, respectively, are indicated by arrows and can be discriminated by the widespread (GA)10 hybridization pattern on the long arms. Scale bars: 5 µm.
Figure 7.
Figure 7.
An example of globe artichoke pairing of homologous chromosomes based on (GA)10 labelling pattern. Scale bar: 5 µm.
Figure 8.
Figure 8.
Ideogram with molecular characterization of cardoon and globe artichoke complement. Ideogram showing chromosome morphology (in black) and the (CA)10, (GA)10 di-nucleotides (red) and pTa71 sequence (green) distribution on cardoon (A) and globe artichoke (B) complement. The two di-nucleotides localize at similar chromosome regions. Scale bar: 5 µm.

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References

    1. Abramoff MD, Magalhaes PJ, Ram SJ. (2004) Image Processing with ImageJ. Biophotonics International 11: 36–42.
    1. Andras SC, Hartman TPV, Marshall JA, Marchant R, Power JB, Cocking EC, Davey MR. (1999) A Drop-spreading Technique to Produce Cytoplasm-free Mitotic Preparations from Plants with Small Chromosomes. Chromosome Research 7: 641–647. doi: 10.1023/A:1009288119851 - DOI - PubMed
    1. Chester M, Leitch AR, Soltis PS, Soltis DE. (2010) Review of the application of modern cytogenetic methods (FISH/GISH) to the study of reticulation (Polyploidy/Hybridization). Gene 1(2): 166–192. doi: 10.3390/genes1010166 - DOI - PMC - PubMed
    1. Cuadrado A, Cardoso M, Jouve N. (2008) Increasing the physical markers of wheat chromosomes using SSRs as FISH probes. Genome 51: 809–815. doi: 10.1139/G08-065 - DOI - PubMed
    1. Doležel J, Binarova P, Lucretti S. (1989) Analysis of nuclear-DNA content in plant-cells by flow-cytometry. Biologia Plantarum 31: 113–120. doi: 10.1007/BF02907241 - DOI

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