The master's thesis was designed with the aim of establishing a protocol for the transformation of four hop varieties, 'Celeia', 'Styrian Eureka', 'Styrian Kolibri' and 'Wye target', for targeted genome editing using CRISPR/Cas9 technology. We tested three different culture media, which varied in hormone content and concentration, and three types of explants: petiole, internode, and leaf explants. The criteria for determining the best combinations were regeneration success, observing callus formation, green callus development, and shoot formation. In the search for the best combination of cultivar, culture medium, and explant type, we observed differences among the cultivars, media, and explant types. For most cultivars, the most suitable medium was the one supplemented with 1 mg/L TDZ and 0.025 mg/L IAA, and among the explants, the internodal explant was the most responsive. For further hop transformation experiments, we excluded the leaf explants due to their poor regeneration and used petiole and internode explants instead. For the transformation, we selected two medium-explant combinations for each variety and introduced the selected transgenes ZsGreen, hptII, and Cas9 on the plasmid pHSE-Zsg-sgHlPDS1 into hop cells using the bacterium Agrobacterium tumefaciens. During explant regeneration, we monitored transformation success using an epifluorescence microscope. To verify the presence of transgenes, we performed PCR analysis and agarose gel electrophoresis. Among all the transgenes, ZsGreen was the most frequently detected in the tested samples, with a 90 % presence rate. The Cas9 transgene was detected in 80 % of the samples, and hptII was present in 65 % of the samples. A comparison of regeneration rates before and after transformation showed a clear difference, as explants regenerated much less successfully after transformation. This study confirms that the success of transformation and regeneration is highly dependent on the specific combination of cultivar, culture medium, and explant type.
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