Manipulation of Gene Activity in the Regenerative Model Sea Anemone, Nematostella vectensis

Autor:	Eric M. Hill, Cheng-Yi Chen, Florencia del Viso, Lacey R. Ellington, Shuonan He, Ahmet Karabulut, Ariel Paulson, Matthew C. Gibson
Rok vydání:	2022
Předmět:	Genome Sea Anemones Mutagenesis Gene Knockdown Techniques Animals RNA Small Interfering Research Article
Zdroj:	Methods Mol Biol Methods in Molecular Biology ISBN: 9781071621714
ISSN:	1940-6029
Popis:	With a surprisingly complex genome and an ever-expanding genetic toolkit, the sea anemone Nematostella vectensis has become a powerful model system for the study of both development and whole-body regeneration. Here we provide the most current protocols for short-hairpin RNA (shRNA)-mediated gene knockdown and CRISPR/Cas9-targeted mutagenesis in this system. We further show that a simple Klenow reaction followed by in vitro transcription allows for the production of gene-specific shRNAs and single guide RNAs (sgRNAs) in a fast, affordable, and readily scalable manner. Together, shRNAknockdown and CRISPR/Cas9-targeted mutagenesis allow for rapid screens of gene function as well as the production of stable mutant lines that enable functional genetic analysis throughout the Nematostella life cycle.
Databáze:	OpenAIRE
Externí odkaz:	https://explore.openaire.eu/search/publication?articleId=doi_dedup___::871a7842a39d66f018e2d2895b0471da https://pubmed.ncbi.nlm.nih.gov/35359322 Zobrazit plný text záznamu