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Intramolecular ligation method (iLIME) for pre-miRNA quantification and sequencing
Hairpin-containing pre-miRNAs, produced from pri-miRNAs, are precursors of miRNAs (microRNAs) that play essential roles in gene expression and various human diseases. Current qPCR-based methods used to quantify pre-miRNAs are not effective to discriminate between pre-miRNAs and their parental pri-mi...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Cold Spring Harbor Laboratory Press
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9202589/ https://www.ncbi.nlm.nih.gov/pubmed/35487691 http://dx.doi.org/10.1261/rna.079101.122 |
Sumario: | Hairpin-containing pre-miRNAs, produced from pri-miRNAs, are precursors of miRNAs (microRNAs) that play essential roles in gene expression and various human diseases. Current qPCR-based methods used to quantify pre-miRNAs are not effective to discriminate between pre-miRNAs and their parental pri-miRNAs. Here, we developed the intramolecular ligation method (iLIME) to quantify and sequence pre-miRNAs specifically. This method utilizes T4 RNA ligase 1 to convert pre-miRNAs into circularized RNAs, allowing us to design PCR primers to quantify pre-miRNAs, but not their parental pri-miRNAs. In addition, the iLIME also enables us to sequence the ends of pre-miRNAs using next-generation sequencing. Therefore, this method offers a simple and effective way to quantify and sequence pre-miRNAs, so it will be highly beneficial for investigating pre-miRNAs when addressing research questions and medical applications. |
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