DLO Hi-C染色体构象捕获技术
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摘要
基因组三维结构在DNA复制、DNA损伤修复、基因转录调控中扮演着重要的角色。Hi-C是目前研究基因组三维结构的主要手段之一,但是存在背景噪音大、试验成本高、试验流程繁琐,缺乏对随机连接噪音的评价等缺陷,因而限制了三维基因组学的发展。为此,我们开发了一种简单经济的染色体构象捕获技术,即DLO Hi-C(digestion-ligation-only Hi-C)。该技术去掉了生物素标记的步骤,只需要2轮简单的酶切酶连反应即可构建高质量的DLO Hi-C测序文库。为了评价文库中的随机连接噪音的比例,我们在文库构建步骤中加入了噪音评价的步骤。研究结果显示,与当前的in situ Hi-C等方法比较,我们的DLO Hi-C试验时间短、试验成本低、测序成本低,有更多的有效交互数据。将DLO Hi-C应用于肿瘤细胞系,我们找到了已知和新的基因组结构变异。所以,我们希望DLO Hi-C技术将对研究染色体的三维构象、基因的转录调控及基因组的组装有重要的促进作用。
The 3D structure of the genome has an important role in DNA replication,DNA damage repair,and transcriptional regulation. However,high background noise,high cost,and a lack of straightforward noise evaluation in the Hi-C methods impede the advancement of 3D genomic research. Here we develop a simple digestion-ligation-only Hi-C(DLO Hi-C) technology to explore the 3D landscape of the genome. This method requires only two rounds of digestion and ligation,without the need for biotin labeling.Notably,random ligation could be quickly evaluated in an early quality-control step before sequencing. Our results show that DLO Hi-C experiment has a shorter time,lower experimental cost,lower sequencing cost,and more effective interaction data than current in situ Hi-C methods. Applying DLO Hi-C to tumor cell lines,we find known and new genomic translocations. This technology may facilitate investigation of genomic organization,gene regulation,and genome assembly.
The 3D structure of the genome has an important role in DNA replication,DNA damage repair,and transcriptional regulation. However,high background noise,high cost,and a lack of straightforward noise evaluation in the Hi-C methods impede the advancement of 3D genomic research. Here we develop a simple digestion-ligation-only Hi-C(DLO Hi-C) technology to explore the 3D landscape of the genome. This method requires only two rounds of digestion and ligation,without the need for biotin labeling.Notably,random ligation could be quickly evaluated in an early quality-control step before sequencing. Our results show that DLO Hi-C experiment has a shorter time,lower experimental cost,lower sequencing cost,and more effective interaction data than current in situ Hi-C methods. Applying DLO Hi-C to tumor cell lines,we find known and new genomic translocations. This technology may facilitate investigation of genomic organization,gene regulation,and genome assembly.
引文
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[2]Dekker J,Belmont A S,Guttman M,et al.The 4D nucleome project[J].Nature,2017,549(7671):219-226.
[3]李国亮,阮一骏,谷瑞升,等.起航三维基因组学研究[J].科学通报,2014,59(13):1165-1172.Li G L,Ruan Y J,Gu R S,et al.Emergence of 3D genomics[J].Chinese Science Bulletin,2014,59(13):1165-1172(in Chinese with English abstract).
[4]Lieberman-Aiden E,Dekker J.Comprehensive mapping of long-range interactions reveals folding principles of the human genome[J].Science,2009,326(5905):289.
[5]Dixon J R,Jung I,Selvaraj S,et al.Chromatin architecture reorganization during stem cell differentiation[J].Nature,2015,518(7539):331-336.
[6]Ke Y,Yu Y,Chen X,et al.3D chromatin structures of mature gametes and structural reprogramming during mammalian embryogenesis[J].Cell,2017,170(2):367-381.
[7]Lin D,Hong P,Zhang S,et al.Digestion-ligation-only Hi-C is an efficient and cost-effective method for chromosome conformation capture[J].Nature Genetics,2018,50(5):754-763.
[8]Cost G J.Enzymatic ligation assisted by nucleases:simultaneous ligation and digestion promote the ordered assembly of DNA[J].Nature Protocols,2007,2(9):2198-2202.