Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9

Peiwu Qin, Mahmut Parlak, Cem Kuscu, Jigar Bandaria, Mustafa Mir, Karol Szlachta, Ritambhara Singh, Xavier Darzacq, Ahmet Yildiz, Mazhar Adli

Research output: Contribution to journalArticle

48 Citations (Scopus)

Abstract

Imaging chromatin dynamics is crucial to understand genome organization and its role in transcriptional regulation. Recently, the RNA-guidable feature of CRISPR-Cas9 has been utilized for imaging of chromatin within live cells. However, these methods are mostly applicable to highly repetitive regions, whereas imaging regions with low or no repeats remains as a challenge. To address this challenge, we design single-guide RNAs (sgRNAs) integrated with up to 16 MS2 binding motifs to enable robust fluorescent signal amplification. These engineered sgRNAs enable multicolour labelling of low-repeat-containing regions using a single sgRNA and of non-repetitive regions with as few as four unique sgRNAs. We achieve tracking of native chromatin loci throughout the cell cycle and determine differential positioning of transcriptionally active and inactive regions in the nucleus. These results demonstrate the feasibility of our approach to monitor the position and dynamics of both repetitive and non-repetitive genomic regions in live cells.

Original languageEnglish (US)
Article number14725
JournalNature Communications
Volume8
DOIs
StatePublished - Mar 14 2017
Externally publishedYes

Fingerprint

Clustered Regularly Interspaced Short Palindromic Repeats
Guide RNA
chromatin
chromosomes
loci
Chromosomes
Chromatin
Imaging techniques
cells
genome
Nucleic Acid Repetitive Sequences
Labeling
positioning
marking
Amplification
Cell Cycle
Genes
Cells
Genome
RNA

All Science Journal Classification (ASJC) codes

  • Chemistry(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Physics and Astronomy(all)

Cite this

Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9. / Qin, Peiwu; Parlak, Mahmut; Kuscu, Cem; Bandaria, Jigar; Mir, Mustafa; Szlachta, Karol; Singh, Ritambhara; Darzacq, Xavier; Yildiz, Ahmet; Adli, Mazhar.

In: Nature Communications, Vol. 8, 14725, 14.03.2017.

Research output: Contribution to journalArticle

Qin, P, Parlak, M, Kuscu, C, Bandaria, J, Mir, M, Szlachta, K, Singh, R, Darzacq, X, Yildiz, A & Adli, M 2017, 'Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9', Nature Communications, vol. 8, 14725. https://doi.org/10.1038/ncomms14725
Qin, Peiwu ; Parlak, Mahmut ; Kuscu, Cem ; Bandaria, Jigar ; Mir, Mustafa ; Szlachta, Karol ; Singh, Ritambhara ; Darzacq, Xavier ; Yildiz, Ahmet ; Adli, Mazhar. / Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9. In: Nature Communications. 2017 ; Vol. 8.
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