Determination of nuclear DNA content, ploidy, and FISH location of ribosomal DNA in Hibiscus hamabo

Sakhanokho, Hamidou F.; Islam‐Faridi, Nurul; Babiker, Ebrahiem M.; Nelson, C. Dana; Stringer, Stephen J.; Adamczyk, John J.

doi:10.1016/j.scienta.2019.109167

Cited by 17 publications

(24 citation statements)

References 27 publications

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“…It is often difficult to obtain well-separated chromosome spreads with full chromosome complements, to accurately determine a species' chromosome number (and/or ploidy), especially from plants with a high number of small chromosomes (~ 50 and higher). Our modified enzymatic digestion of protoplast technique 40 works well to prepare chromosome spreads from plant species with a higher number of small chromosomes like A. digitata (this report) or Hibiscus hamabo 41 . A key feature of this method is allowing the chromosomes to spread naturally on the glass slides without squashing them with cover slips ( for CMA3 banding, and nine and ten species excluding A. digitata for 45S rDNA and 5S rDNA, respectively, with FISH 35 .…”

Section: Discussionmentioning

confidence: 91%

“…Data from interphase nuclei provide an upwardly biased count of the number of tandemly repetitive DNA loci such as 45S rDNA. It has been reported that the interphase nuclei cannot be used to determine the 45S rDNA loci number due to their decondensed nature 41 , 43 , 48 , 61 and this is supported by genome sequencing where rDNA loci are often missing from genome assemblies as they cannot be correctly assembled 47 . We suggest this is also the case for prophase and pro-metaphase when the 45S rDNA is massively large like in A. digitata .…”

Section: Discussionmentioning

confidence: 99%

“…Flow cytometry was performed as described by Sakhanokho et al 41 with minor modifications to the nuclei staining solution and procedure. In this study, we stained A. digitata nuclei with propidium iodide (PI) for analysis with a BD Accuri C6 flow cytometer and a BD Accuri C6 software version 1.0.264.21 (BD BioSciences, Ann Arbor, MI).…”

Section: Methodsmentioning

confidence: 99%

“…S8 , Seedling-1 and Seedling-2) were used for root tip chromosome spreads and processed separately. Chromosome spreads were prepared following procedures previously described 40 , 41 with some modifications. Actively growing root tips about 1.0 cm long were harvested and immediately pre-treated with 2 mM (0.036% (w/v)) 8-hydroxyquilonine for 4.0 h in the dark at room temperature (RT, 22–24 °C), rinsed with ddH 2 O and then fixed in 4:1 (95% ethanol (EtOH): glacial acetic acid (GAA)) and stored at RT over-night before processing for enzyme digestion for chromosome spread.…”

Section: Methodsmentioning

confidence: 99%

“…Given these conflicting results for chromosome number and genome size of A. digitata , we decided to reevaluate these parameters using updated techniques. For chromosome number we used a root tip protoplast technique for chromosome spreading 40 , 41 and for genome size we used flow cytometry. Chromosome banding using Chromamycin A3 (CMA3) involving A. digitata has been reported 35 , but no reports on fluorescence in situ hybridization (FISH) using rDNA or other probes in this species are available.…”

Section: Introductionmentioning

confidence: 99%

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New chromosome number and cyto-molecular characterization of the African Baobab (Adansonia digitata L.) - “The Tree of Life”

Islam‐Faridi

Sakhanokho

Nelson

2020

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new chromosome number and cyto-molecular characterization of the African Baobab (Adansonia digitata L.)-"the tree of Life" nurul islam-faridi 1,5* , Hamidou F. Sakhanokho 2,5 & c. Dana nelson 3,4 the African baobab (Adansonia digitata L.), also referred to as the "Tree of Life", is a majestic, longlived and multipurpose tree of sub-Saharan Africa. Internationally, a growing demand for baobab products in the food, pharmaceutical and cosmetics industries has been observed. Considering this, there is a need for scientific information on the genetics and breeding of A. digitata, including cytogenetics, genetic diversity and reproductive biology. The objectives of our cytogenetic research were to determine the genome size, chromosome number, and organization of ribosomal DNA (45S and 5SrDNA) of A. digitata. Flow cytometry analysis revealed a 2C-DNA value of 3.8 ± 0.6 pg (1Cx monoploid genome size 919.1 ± 62.9 Mbp). Using our improved chromosome preparation technique, we were able to unequivocally count the chromosomes resulting in 2n = 4x = 168, a revised chromosome number for A. digitata. Fluorescent in situ hybridization (FISH) analysis revealed two massively large variants of 45S rDNA and their corresponding nucleolus organizer regions (NOR). The NOR variants were about two to four times larger than the main body of their respective chromosomes. To our knowledge, this is the first report of this phenomenon in a plant species. Furthermore, we found that FISH analysis using the Arabidopsis-type telomere repeat sequence probe clarified and confirmed the new chromosome number and characterized the 45S rDNA structural organization. The African baobab (Adansonia digitata L.) is the largest and best known of the eight Adansonia species and reported to be native to mainland Africa 1-3. It is widespread throughout the hot, drier regions of tropical Africa, extending from northern Tanzania and Namibia to Ethiopia, Sudan, and the northern fringes of the Sahara 1. Adansonia digitata is often referred to by its vernacular name, baobab, which is believed to originate from the Arabic word "buhibab" or "fruit with many seeds" 4,5. The genus Adansonia is named after Michel Adanson who brought baobab seeds to Paris in 1754 and who was the first person to provide a comprehensive description and drawing of the tree after his visit to Senegal 5,6. Despite having hermaphrodite flowers, A. digitata is mainly selfincompatible 7. This majestic tree, known as the "Tree of Life", is deciduous, reaching up to 20-25 m in height and 20 m in trunk diameter 1,2,8 and living for hundreds or even thousands of years 9. A recent radiocarbon dating 10 study estimated the age of the Glencoe baobab tree located in the Limpopo Province of South Africa to be about 1,800 years 11. The baobab is a multipurpose tree that is extensively used for both its nutritional and medicinal values. Virtually, every part of the baobab tree is useful, and the "Tree of Life" lives up to its name as it is reported to have

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Section: Discussionmentioning

confidence: 91%