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J Exp Bot. 2016 Nov;67(21):6101-6110. Epub 2016 Oct 6.

Visualization of specific repetitive genomic sequences with fluorescent TALEs in Arabidopsis thaliana.

Author information

1
Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Noda, Chiba 278-8510, Japan.
2
Graduate School of Science, Kyoto University, Kyoto 606-8502, Japan.
3
PRESTO, JST, Saitama 332-0012, Japan.
4
Institute of Transformative Bio-Molecules, Nagoya University, Nagoya 464-8601, Japan.
5
Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima 770-8513, Japan.
6
Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Noda, Chiba 278-8510, Japan sachi@rs.tus.ac.jp.

Abstract

Live imaging of the dynamics of nuclear organization provides the opportunity to uncover the mechanisms responsible for four-dimensional genome architecture. Here, we describe the use of fluorescent protein (FP) fusions of transcription activator-like effectors (TALEs) to visualize endogenous genomic sequences in Arabidopsis thaliana. The ability to engineer sequence-specific TALEs permits the investigation of precise genomic sequences. We could detect TALE-FP signals associated with centromeric, telomeric, and rDNA repeats and the signal distribution was consistent with that observed by fluorescent in situ hybridization. TALE-FPs are advantageous because they permit the observation of intact tissues. We used our TALE-FP method to investigate the nuclei of several multicellular plant tissues including roots, hypocotyls, leaves, and flowers. Because TALE-FPs permit live-cell imaging, we successfully observed the temporal dynamics of centromeres and telomeres in plant organs. Fusing TALEs to multimeric FPs enhanced the signal intensity when observing telomeres. We found that the mobility of telomeres was different in sub-nuclear regions. Transgenic plants stably expressing TALE-FPs will provide new insights into chromatin organization and dynamics in multicellular organisms.

KEYWORDS:

Centromere; chromatin; fluorescent protein; genome editing; live cell imaging; rDNA; telomere; transcription activator-like effector.

PMID:
27811079
PMCID:
PMC5100022
DOI:
10.1093/jxb/erw371
[Indexed for MEDLINE]
Free PMC Article

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