Cytogenetic and cytometric analyses in artificial intercytotypic hybrids of the emergent orchid model species Erycina pusilla

Abstract

Erycina pusilla is considered a potential model organism for orchids, because of several advantageous features, such as short juvenile period, low chromosome number and all year round blooming. Two different chromosome numbers (2n = 10 and 2n = 12) are reported for E. pusilla, which suggests two cytotypes. To reveal the genome homology between these two cytotypes, we generated hybrids from the intercytotypic reciprocal crosses (2n = 10 9 2n = 12 and 2n = 12 9 2n = 10), and applied 4 0 ,6-diamidino-2-phenylindole staining, genomic in situ hybridization and flow cytometry for genomic and cytogenetic analysis. The parental genomes showed high similarity both in genomic composition and content. The hybrids displayed a chromosome number of 2n = 11 in mitotic cells. Moreover, five bivalents and one univalent were observed at meiotic metaphase I stage. We observed meiotic synaptic behavior and found homeologous pairing with unpaired loops between parental chromosome pairing segments. These results demonstrated that chromosome rearrangement events have occurred between parental cytotypes during evolution. This study also illustrated the genome homology and homeologous pairing at pachytene phase, indicating that the chromosome number variation of two cytotypes mainly resulted from chromosome rearrangements, not changes in genomic constitution.

Flow cytometry – a modern method for exploring genome size and nuclear DNA synthesis in horticultural and medicinal plant species

“…Their study confirmed earlier observations by Johnston et al (2005), who, after measuring the DNA content of 34 Brassicaceae species and sequencing the ITS region (internal transcribed spacer region of DNA between the small and large ribosomal RNA genes) for 23 taxa, had found that despite the evidence of species with decreases and increases in GS, including polyploidization, the evolution of this family's GS had a conservative nature. Polyploidy, and also dysploidy (an increase or decrease in chromosome number as a result of structural rearrangements) as mechanisms of genome evolution have been suggested for Ocimum, Orchidaceae, Bromeliaceae, Colchicum, and Iridaceae (Bory et al, 2008;Carović-Stanko et al, 2010;Rupp et al, 2010;Trávníček et al, 2011;Moraes et al, 2012Moraes et al, , 2015Jersáková et al, 2013;Fridlender et al, 2014;Gitaí et al, 2014;Yeh et al, 2015).…”

Two reported cytotypes of the emergent orchid model species Erycina pusilla are two different species

“…There is a series of multiple chromosome rearrangements required to generate the differences between the two karyotypes. Our previous result from Yeh et al (2015) as well as our subsequent efforts also showed no evidence so far for fertile F1 hybrids between the parents with the two different karyotypes. The sterile F1 hybrids and our findings in the study support the hypothesis that these are two separate species, not just cytotypes.…”

“…The two cytotypes of E. pusilla were collected from Ecuador (2n = 10) and from Surinam (2n = 12) and their specimens were deposited in the herbarium of Taiwan's National Museum of Natural Science (Yeh et al 2015). All plants were maintained and cultured on half-strength Murashige and Skoog medium (Murashige and Skoog 1962), and cultivated in growth chambers under long-day conditions (16-h photoperiod) at 22°C.…”

“…For example, the basic chromosome number of Paphiopedilum was proposed as 2n = 26, and the chromosome number variation under the genus ranged from 2n = 26-42 caused by a Robertsonian centric fission mechanism (Jones 1998;Lee et al 2011). It has been proposed that the change in chromosome number is from 12 to 10, even initially from 14, in this species (Yeh et al 2015) because E. pusilla is in the Cymbidioid group, in which the most probable basic chromosome number is x = 7 (Félix and Guerra 2000. In this study, we have demonstrated divergent chromosome profiles between two karyotypes that are mostly likely due to multiple steps in chromosome rearrangements.…”

“…However, little cytogenetic information has been reported: only a chromosome count and conventional karyotype analysis has been carried out in this orchid. In Yeh et al (2015) the two 2n = 10 and 2n = 12 cytotypes, collected from two South American countries, demonstrated high similarity both in genomic composition and DNA amount based on the results of genomic in situ hybridization and flow cytometry. In addition, analyses of meiotic prophase of the 2n = 10/12 heterozygotes from reciprocal crosses revealed incomplete pairing, with unpaired loops between parental homoeologous chromosome segments.…”