A Nonsequencing Approach for the Rapid Detection of RNA Editing

Ruchika, Toshifumi Tshukahara, Manish Biyani

研究成果: ジャーナルへの寄稿学術誌査読

抄録

RNA editing is a process that leads to posttranscriptional sequence alterations in RNAs. Detection and quantification of RNA editing rely mainly on Sanger sequencing and RNA sequencing techniques. However, these methods can be costly and timeconsuming. In this protocol, a portable microtemperature gradient gel electrophoresis (µTGGE) system is used as a nonsequencing approach for the rapid detection of RNA editing. The process is based on the principle of electrophoresis, which uses high temperatures to denature nucleic acid samples as they move across a polyacrylamide gel. Across a range of temperatures, a DNA fragment forms a gradient of fully double-stranded DNA to partially separated strands and then to entirely separated single-stranded DNA. RNA-edited sites with distinct nucleotide bases produce different melting profiles in µTGGE analyses. We used the µTGGE-based approach to characterize the differences between the melting profiles of four edited RNA fragments and their corresponding nonedited (wild-type) fragments. Pattern Similarity Scores (PaSSs) were calculated by comparing the band patterns produced by the edited and nonedited RNAs and were used to assess the reproducibility of the method. Overall, the platform described here enables the detection of even single base mutations in RNAs in a straightforward, simple, and cost-effective manner. It is anticipated that this analysis tool will aid new molecular biology findings.

本文言語英語
論文番号e63591
ジャーナルJournal of Visualized Experiments
2022
182
DOI
出版ステータス出版済み - 4月 2022
外部発表はい

!!!All Science Journal Classification (ASJC) codes

  • 神経科学(全般)
  • 化学工学(全般)
  • 生化学、遺伝学、分子生物学(全般)
  • 免疫学および微生物学(全般)

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