Hyperexpansion of wheat chromosomes sorted by flow cytometry

Despite remarkable recent progress in the analysis of plant genome organization and chromosome structure, there is a need for methods enabling DNA sequences to be mapped by fluorescence in situ hybridization (FISH) at high spatial resolution. We sorted mitotic metaphase chromosomes of wheat by flow...

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Bibliographic Details
Published in:Genes & Genetic Systems 2014/08/01, Vol.89(4), pp.181-185
Main Authors: Endo, Takashi R., Kubaláková, Marie, Vrána, Jan, Doležel, Jaroslav
Format: Article
Language:English
Subjects:
chromosome
Chromosomes, Plant - genetics
Chromosomes, Plant - ultrastructure
FISH
Flow Cytometry
flow sorting
hyperexpansion
In Situ Hybridization, Fluorescence
Triticum - genetics
Triticum aestivum
wheat
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Summary:Despite remarkable recent progress in the analysis of plant genome organization and chromosome structure, there is a need for methods enabling DNA sequences to be mapped by fluorescence in situ hybridization (FISH) at high spatial resolution. We sorted mitotic metaphase chromosomes of wheat by flow cytometry and observed the occurrence of hyperexpanded chromosomes among them. However, this phenomenon was not reproducible in subsequent experiments. An investigation into the procedures of flow cytometry revealed that the hyperexpansion of chromosomes became reproducible when the concentration of formaldehyde used in sample fixation was reduced. We conducted FISH analysis with 45S rDNA, 5S rDNA and wheat centromeric repeat sequences as probes on flow-sorted chromosomes and also on chromosomes from squash preparations. We measured the length of chromosomes 1B and 6B, identified by FISH. On average, the hyperexpanded 1B and 6B chromosomes were 7.26 and 7.53 times longer, respectively, than the same chromosomes from the squash preparations. The most stretched 1B and 6B chromosomes both exceeded 100 micrometers.
ISSN:1341-7568
1880-5779
DOI:10.1266/ggs.89.181