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Bokkapitel2021Vetenskapligt granskad

Protoplast-Based Method for Genome Editing in Tetraploid Potato

Nicolia, Alessandro; Fält, Ann-Sofie; Hofvander, Per; Andersson, Mariette

Sammanfattning

The cultivated potato is tetraploid with four probably equivalent loci for each gene. A potato variety is furthermore commonly genetically heterogeneous and selected based on a beneficial genetic context which is maintained by clonal propagation. When introducing genetic changes by genome editing it is then desirable to achieve edits in all four loci for a certain gene target. This is in order to avoid crosses to achieve homozygosity for edited gene loci and at the same time reduce risk of inbreeding depression. In such a context transient transfection of protoplasts for the introduction of mutations, avoiding stable insertion of foreign DNA, would be very attractive. The protocol of this chapter has been shown to be applicable for the introduction of mutations by DNA vectors containing expression cassettes of TALEN, Cas9, and Cas9 deaminase fusions together with sgRNA expression cassettes on either single or separate vectors. Furthermore, the protoplast-based system has been shown to work very efficiently for mutations introduced by in vitro-produced and transfected RNP (ribonucleoprotein) complexes.

Nyckelord

Potato; Genome editing; Targeted mutagenesis; TALEN; CRISPR-Cas9; DNA vector; RNP

Publicerad i

Methods in Molecular Biology
2021, nummer: 2264, sidor: 177-186 Titel: Crop Breeding : Genetic Improvement Methods
ISBN: 978-1-0716-1200-2, eISBN: 978-1-0716-1201-9Utgivare: Humana
DOI: 10.1007/978-1-0716-1201-9_12