An improved method for inducing prometaphase chromosomes in plants

Setiawan, Agus Budi and Teo, Chee How and Kikuchi, Shinji and Sassa, Hidenori and Koba, Takato (2018) An improved method for inducing prometaphase chromosomes in plants. Molecular Cytogenetics, 11 (1). p. 32. ISSN 1755-8166, DOI

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Background: Detailed karyotyping using metaphase chromosomes in melon (Cucumis melo L.) remains a challenge because of their small chromosome sizes and poor stainability. Prometaphase chromosomes, which are two times longer and loosely condensed, provide a significantly better resolution for fluorescence in situ hybridization (FISH) than metaphase chromosomes. However, suitable method for acquiring prometaphase chromosomes in melon have been poorly investigated. Results: In this study, a modified Carnoy's solution II (MC II) [6:3:1 (v/v) ethanol: acetic acid: chloroform] was used as a pretreatment solution to obtain prometaphase chromosomes. We demonstrated that the prometaphase chromosomes obtained using the MC II method are excellent for karyotyping and FISH analysis. We also observed that a combination of MC II and the modified air dry (ADI) method provides a satisfactory meiotic pachytene chromosome preparation with reduced cytoplasmic background and clear chromatin spreads. Moreover, we demonstrated that pachytene and prometaphase chromosomes of melon and Abelia × grandiflora generate significantly better FISH images when prepared using the method described. We confirmed, for the first time, that Abelia × grandiflora has pairs of both strong and weak 45S ribosomal DNA signals on the short arms of their metaphase chromosomes. Conclusion: The MC II and ADI method are simple and effective for acquiring prometaphase and pachytene chromosomes with reduced cytoplasm background in plants. Our methods provide high-resolution FISH images that can help accelerate molecular cytogenetic research in plants.

Item Type: Article
Funders: Grants-in-Aid for Scientific Research (No. 16K07588) from the Ministry of Education, Culture, Sports, Science and Technology, Japan
Uncontrolled Keywords: Abelia × grandiflora; Chloroform; Cucumis melo; FISH; Pachytene; Prometaphase
Subjects: Q Science > Q Science (General)
Q Science > QH Natural history
Divisions: Deputy Vice Chancellor (Research & Innovation) Office > Centre for Research in Biotechnology for Agriculture
Depositing User: Ms. Juhaida Abd Rahim
Date Deposited: 19 Mar 2019 02:20
Last Modified: 19 Mar 2019 02:20

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