title: Fluorescence in situ hybridization on plant extended chromatin DNA fibers for single-copy and repetitive DNA sequences.
abstract: The compactness of plant chromosomes and the structure of the plant cell wall and cytoplasm provide a great obstacle to fluorescence in situ hybridization (FISH) for single-copy or low-copy DNA sequences. Consequently, many new methods for improving spatial resolution via chromosomal stretching have been employed to overcome this technical challenge. In this article, a technique for extracting cell-wall free nuclei at mitotic interphase, then using these nuclei to prepare extended DNA fibers (EDFs) by the method of a receding interface, whereby slide-mounted chromatin produces EDFs in concert with gravity-assisted buffer flow, was adopted as a result of the low frequency of EDF damage produced by this procedure. To examine the quality of these EDFs, we used single-copy gene encoding S-locus receptor kinase and multi-copy 5S rDNA (ribosomal DNA) as probes. The resulting EDFs proved suitable for high-resolution FISH mapping for repetitive DNA sequences, and the localization of a single-copy locus.
authors.affiliation: College of Agronomy and Biotechnology, Southwest University, Chongqing, China.
authors.forename: Kun
authors.initials: K
authors.lastname: Yang
authors.forename: Hecui
authors.initials: H
authors.lastname: Zhang
authors.forename: Richard
authors.initials: R
authors.lastname: Converse
authors.forename: Yong
authors.initials: Y
authors.lastname: Wang
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Rong
authors.forename: Zhigang
authors.initials: Z
authors.lastname: Wu
authors.forename: Bing
authors.initials: B
authors.lastname: Luo
authors.forename: Liyan
authors.initials: L
authors.lastname: Xue
authors.forename: Li
authors.initials: L
authors.lastname: Jian
authors.forename: Liquan
authors.initials: L
authors.lastname: Zhu
authors.forename: Xiaojia
authors.initials: X
authors.lastname: Wang
mesh_terms: D001937:Brassica / Q000235:genetics; D002467:Cell Nucleus / Q000235:genetics; D002473:Cell Wall / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D002874:Chromosome Mapping / Q000379:methods*; D032461:Chromosomes, Plant / Q000235:genetics; D015342:DNA Probes / Q000235:genetics / Q000378:metabolism; D018744:DNA, Plant / Q000235:genetics*; D035264:Flowers / Q000235:genetics; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D007399:Interphase; D008677:Metaphase; D049471:Pachytene Stage; D010940:Plant Proteins / Q000235:genetics; D018517:Plant Roots / Q000235:genetics; D011494:Protein Kinases / Q000235:genetics; D012341:RNA, Ribosomal, 5S / Q000235:genetics / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid
chemical_list: D002843:Chromatin; D015342:DNA Probes; D018744:DNA, Plant; D010940:Plant Proteins; D012341:RNA, Ribosomal, 5S; D011494:Protein Kinases; C079832:S-receptor kinase