Yang Zhang, Steentoft Catharina, Hauge Camilla, Hansen Lars, Thomsen Allan Lind, Niola Francesco, Vester-Christensen Malene B, Frödin Morten, Clausen Henrik, Wandall Hans H, Bennett Eric P
Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, 2200 Copenhagen N, Denmark Novo Nordisk Foundation Center for Biosustainability, Danish Technical University, Lyngby, Denmark.
Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, 2200 Copenhagen N, Denmark.
Nucleic Acids Res. 2015 May 19;43(9):e59. doi: 10.1093/nar/gkv126. Epub 2015 Mar 9.
The nuclease-based gene editing tools are rapidly transforming capabilities for altering the genome of cells and organisms with great precision and in high throughput studies. A major limitation in application of precise gene editing lies in lack of sensitive and fast methods to detect and characterize the induced DNA changes. Precise gene editing induces double-stranded DNA breaks that are repaired by error-prone non-homologous end joining leading to introduction of insertions and deletions (indels) at the target site. These indels are often small and difficult and laborious to detect by traditional methods. Here we present a method for fast, sensitive and simple indel detection that accurately defines indel sizes down to ±1 bp. The method coined IDAA for Indel Detection by Amplicon Analysis is based on tri-primer amplicon labelling and DNA capillary electrophoresis detection, and IDAA is amenable for high throughput analysis.
基于核酸酶的基因编辑工具正在迅速改变细胞和生物体基因组的精确改变能力，并可用于高通量研究。精确基因编辑应用中的一个主要限制在于缺乏灵敏且快速的方法来检测和表征诱导的DNA变化。精确基因编辑会诱导双链DNA断裂，这些断裂通过易出错的非同源末端连接进行修复，从而导致在靶位点引入插入和缺失（indel）。这些indel通常很小，用传统方法检测困难且费力。在此，我们提出一种快速、灵敏且简单的indel检测方法，该方法能准确确定低至±1 bp的indel大小。我们将该方法命名为IDAA（通过扩增子分析进行Indel检测），它基于三引物扩增子标记和DNA毛细管电泳检测，并且适用于高通量分析。
