Manipulation of Gene Activity in the Regenerative Model Sea Anemone, Nematostella vectensis.
Methods Mol Biol
; 2450: 437-465, 2022.
Article
em En
| MEDLINE
| ID: mdl-35359322
ABSTRACT
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, shRNA knockdown 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.
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Base de dados:
MEDLINE
Assunto principal:
Anêmonas-do-Mar
Idioma:
En
Ano de publicação:
2022
Tipo de documento:
Article