Increasing of Amylopectin in Tetraploid Potato Desiree Cultivar (Solanum tuberosum L.) Via CRISPR/Cas9

Abstract

Potato tuber starch characteristic is influenced by the ratio of amylose to amylopectin, which is affected by the granule-bound starch synthase (GBSS) gene. GBSS gene expression was decreased in the leaves of the Desiree potato variety utilizing genome editing using CRISPR-Cas9. Constructs encoding Cas9 gene and sgRNAs targeting GBSS gene were inserted into plant leaves using Agrobacterium-mediated transformation delivery. The results obtained lines with mutations in GBSS genes accounting for 21% of regenerated shoots. The identification of mutations within one base pair of the used guide sequence provided further evidence of the considerable similarity between the target region around the protospacer adjacent motif (PAM) position and the used guide sequence. Transforming DNA into potato leaves produced mutants that lacked the Cas9 gene. Using microscopic inspection of iodine-stained starch granules, the increase of amylopectin in the starch granules of editing potato tubers was evaluated.

Keywords

• crispr-cas9
• amylopectin
• granule-bound starch synthase
• sgrnas
• agrobacterium- mediated transformation