Long-term dual-color tracking of genomic loci by modified sgRNAs of the   CRISPR/Cas9 system

doi:10.1093/nar/gkw066

CORC > 北京大学 > 生命科学学院

	Long-term dual-color tracking of genomic loci by modified sgRNAs of the CRISPR/Cas9 system
	Shao, Shipeng ; Zhang, Weiwei ; Hu, Huan ; Xue, Boxin ; Qin, Jinshan ; Sun, Chaoying ; Sun, Yuao ; Wei, Wensheng ; Sun, Yujie
刊名	NUCLEIC ACIDS RESEARCH
	2016
关键词	SINGLE-PARTICLE TRACKING EMBRYONIC STEM-CELLS DNA-SEQUENCES LIVING CELLS CHROMOSOME DYNAMICS CHROMATIN DYNAMICS NUCLEAR LAMINA TARGET DNA GENE RNA
DOI	10.1093/nar/gkw066
英文摘要	Visualization of chromosomal dynamics is important for understanding many fundamental intra-nuclear processes. Efficient and reliable live-cell multicolor labeling of chromosomal loci can realize this goal. However, the current methods are constrained mainly by insufficient labeling throughput, efficiency, flexibility as well as photostability. Here we have developed a new approach to realize dual-color chromosomal loci imaging based on a modified single-guide RNA (sgRNA) of the CRISPR/Cas9 system. The modification of sgRNA was optimized by structure-guided engineering of the original sgRNA, consisting of RNA aptamer insertions that bind fluorescent protein-tagged effectors. By labeling and tracking telomeres, centromeres and genomic loci, we demonstrate that the new approach is easy to implement and enables robust dual-color imaging of genomic elements. Importantly, our data also indicate that the fast exchange rate of RNA aptamer binding effectors makes our sgRNA-based labeling method much more tolerant to photobleaching than the Cas9-based labeling method. This is crucial for continuous, long-term tracking of chromosomal dynamics. Lastly, as our method is complementary to other live-cell genomic labeling systems, it is therefore possible to combine them into a plentiful palette for the study of native chromatin organization and genome ultrastructure dynamics in living cells.; National Science Foundation of China [863 Program] [21573013, 21390412, 31271423, 31327901, SS2015AA020406]; National Science Foundation of China [973 Program] [21390412, 31327901]; National Science Foundation of China [Thousand Youth Talents Program]; SCI(E); PubMed; ARTICLE; sun_yujie@pku.edu.cn; 9; e86; 44
语种	英语
内容类型	期刊论文
源URL	[http://ir.pku.edu.cn/handle/20.500.11897/435041]
专题	生命科学学院
推荐引用方式 GB/T 7714	Shao, Shipeng,Zhang, Weiwei,Hu, Huan,et al. Long-term dual-color tracking of genomic loci by modified sgRNAs of the CRISPR/Cas9 system[J]. NUCLEIC ACIDS RESEARCH,2016.
APA	Shao, Shipeng.,Zhang, Weiwei.,Hu, Huan.,Xue, Boxin.,Qin, Jinshan.,...&Sun, Yujie.(2016).Long-term dual-color tracking of genomic loci by modified sgRNAs of the CRISPR/Cas9 system.NUCLEIC ACIDS RESEARCH.
MLA	Shao, Shipeng,et al."Long-term dual-color tracking of genomic loci by modified sgRNAs of the CRISPR/Cas9 system".NUCLEIC ACIDS RESEARCH (2016).