Live cell imaging of single genomic loci with quantum dot-labeled TALEs.
Live cell imaging of single genomic loci with quantum dot-labeled TALEs.
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使用量子点标记的 TALE 对单个基因组位点进行活细胞成像
DOI：
10.1038/ncomms15318
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发表时间：
2017-05-08
影响因子：
16.6
通讯作者：
Cui Z
中科院分区：
文献类型：
--
作者：
Ma Y;Wang M;Li W;Zhang Z;Zhang X;Tan T;Zhang XE;Cui Z
单个基因组位点通常与特定的细胞功能、遗传疾病或病原性感染相关。活的人类细胞中的单个基因组位点的可视化目前引起了极大的兴趣，但它仍然具有挑战性。在这里，我们描述了一种策略，通过结合转录激活因子样效应物（TALE）与量子点标记技术的单基因组位点的活细胞成像。我们设计并选择了一对特异性靶向HIV-1前病毒DNA序列的TALE，并使用生物正交连接反应用不同颜色的量子点（QD）标记它们。这些QD标记的TALE能够进入细胞核以提供荧光信号来识别单基因位点。基于一对不同颜色的量子点标记的TALE的共定位，我们确定并映射活宿主细胞中人类染色体中的单拷贝HIV-1前病毒基因座。在活细胞中可视化单个基因组位点仍然具有技术挑战性。在这里，作者采用了一种用不同颜色的量子点标记一对转录激活因子样效应子（TALE）的策略，并使用它们来识别活细胞中整合的HIV-1前病毒DNA序列。
Single genomic loci are often related to specific cellular functions, genetic diseases, or pathogenic infections. Visualization of single genomic loci in live human cells is currently of great interest, yet it remains challenging. Here, we describe a strategy for live cell imaging of single genomic loci by combining transcription activator-like effectors (TALEs) with a quantum dot labelling technique. We design and select a pair of TALEs that specifically target HIV-1 proviral DNA sequences, and use bioorthogonal ligation reactions to label them with different colour quantum dots (QDs). These QD-labelled TALEs are able to enter the cell nucleus to provide fluorescent signals to identify single gene loci. Based on the co-localization of the pair of different coloured QD-labelled TALEs, we determine and map single-copy HIV-1 provirus loci in human chromosomes in live host cells. Visualizing single genomic loci in living cells remains technically challenging. Here the authors adopt a strategy of labelling a pair of transcription activator-like effectors (TALEs) with differently coloured quantum dots and use them to identify integrated HIV-1 proviral DNA sequences in living cells.