Mutageneze v Danio rerio pomocí CRISPR technologie

Nickl, Petr

CRISPR/Cas9 technology is currently one of the most important tools of genome engineering. This technology allows a precise site-specific gene editing and such ability was applied to study the role of TALE (TALE - three amino acids loop extension) homeodomain transcription factors during neural crest cells development. The main genes of interest, belonging to sub-family of TALE proteins, are Meis1 transcription factors that are present in the zebrafish genome as two paralogous genes, meis1a and meis1b. Their function was assessed by mutating their DNA-binding domain - homeodomain to abrogate the ability of transcription factor to bind DNA and by that disturb regulatory network, in which Meis1 proteins operate in. Phenotype analysis of mutant fish would reveal a potential role of Meis1 proteins in regulation of neural crest cells development and outline the functional significance of the homeodomain in regulatory operations. To determine the regulatory relationship between meis1a and meis1b genes morpholino-based knock-down of the genes was performed. Preliminary results suggest a dominant role of Meis1b in neural crest cells regulation and importance of its homeodomain. Furthermore, knock-down of Meis1a indicates its contribution to regulation of craniofacial development. However, a detailed...

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