論文

国際誌
2020年9月10日

Rapid, precise quantification of large DNA excisions and inversions by ddPCR.

Scientific reports
  • Hannah L Watry
  • ,
  • Carissa M Feliciano
  • ,
  • Ketrin Gjoni
  • ,
  • Gou Takahashi
  • ,
  • Yuichiro Miyaoka
  • ,
  • Bruce R Conklin
  • ,
  • Luke M Judge

10
1
開始ページ
14896
終了ページ
14896
記述言語
英語
掲載種別
研究論文(学術雑誌)
DOI
10.1038/s41598-020-71742-z

The excision of genomic sequences using paired CRISPR-Cas nucleases is a powerful tool to study gene function, create disease models and holds promise for therapeutic gene editing. However, our understanding of the factors that favor efficient excision is limited by the lack of a rapid, accurate measurement of DNA excision outcomes that is free of amplification bias. Here, we introduce ddXR (droplet digital PCR eXcision Reporter), a method that enables the accurate and sensitive detection of excisions and inversions independent of length. The method can be completed in a few hours without the need for next-generation sequencing. The ddXR method uncovered unexpectedly high rates of large (> 20 kb) excisions and inversions, while also revealing a surprisingly low dependence on linear distance, up to 170 kb. We further modified the method to measure precise repair of excision junctions and allele-specific excision, with important implications for disease modeling and therapeutic gene editing.

リンク情報
DOI
https://doi.org/10.1038/s41598-020-71742-z
PubMed
https://www.ncbi.nlm.nih.gov/pubmed/32913194
PubMed Central
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7483445
ID情報
  • DOI : 10.1038/s41598-020-71742-z
  • PubMed ID : 32913194
  • PubMed Central 記事ID : PMC7483445

エクスポート
BibTeX RIS