@inbook{63575f9843ff4d9cbbb3da71126292fb,
title = "An Efficient CRISPR-Cas9 Method to Knock Out MiRNA Expression in Xenopus Tropicalis",
abstract = "In recent years CRISPR-Cas9 knockouts (KO) have become increasingly utilized to study gene function. MicroRNAs (miRNAs) are short noncoding RNAs, 20–25 nucleotides long, which affect gene expression through posttranscriptional repression. As miRNAs are so small and due to the limitations of known PAM sequences, it is difficult to design CRISPR sgRNAs that reproducibly lead to a KO. We have therefore developed a novel approach using two guide RNAs to effectively “drop out” a miRNA. Validation of efficient CRISPR miRNA KO and phenotype analysis included use of q-RT-PCR and Sanger sequencing. To show specificity of the phenotype, we provide a protocol to use miRNA mimics to rescue the KO phenotype.",
keywords = "CRISPR-Cas9, Knock-out, Micro RNA, Rescue, Xenopus tropicalis",
author = "Godden, {Alice M.} and Marco Antonaci and Wheeler, {Grant N.}",
note = "Funding: This work was supported by UKRI Biotechnology and Biological Sciences Research Council Norwich Research Park Biosciences Doctoral Training Partnership grants to AMG (BB/M011216/1) and the European Union{\textquoteright}s Horizon 2020 research and innovation program under the Marie Sk{\l}odowska-Curie grant agreement no. 860635 to MA.",
year = "2023",
month = jan,
day = "24",
doi = "10.1007/978-1-0716-2982-6_15",
language = "English",
isbn = "978-1-0716-2981-9",
series = "Methods in Molecular Biology",
publisher = "Humana Press",
pages = "231--241",
editor = "Tamas Dalmay",
booktitle = "MicroRNA Detection and Target Identification",
address = "United States",
}