ISHS Acta Horticulturae 1399: XIV International Citrus Congress
Genome editing of IKU1 to obtain citrus seedless fruits |
| Authors: | L. Poles, A. Ciacciulli, H.D. Pappalardo, F. Salonia, G. Distefano, A. Gentile, M. Caruso, S. Larger, M. Pindo, S. La Malfa, C. Licciardello |
| Keywords: | seed, HAIKU, CRISPR/Cas9, mutation, GoldenBraid |
| DOI: | 10.17660/ActaHortic.2024.1399.17 |
| Abstract:
Seedlessness is a desired characteristic in citrus fruits and it is particularly requested by consumers; in the past years many efforts have been done to obtain seedless fruits using conventional breeding techniques, especially in mandarin and mandarin-like cultivars. The use of genome editing and new plant breeding techniques contributes to overcome some limits of conventional breeding and allows the modification of specific traits while maintaining the favourable genetic background of local cultivars. Here, CRISPR/Cas9 genome editing approach was applied to transform seedy citrus cultivars so to try to reduce seed presence by the editing of HAIKU1 (IKU1); this gene codes for a protein that is involved in the development of the seed zygotic tissue and thus in the regulation of the seed size. The construct pIKU-editing_GB was assembled using GoldenBraid 3.0 technology and consists of a dual single guide RNAs (sgRNA1 and sgRNA2) so to be able to induce different types of mutation in IKU1 gene in ‘Carrizo’ citrange. The editing consisted mostly in short insertions and deletions, and in some cases it was possible to recognize larger deletions between the two RNA guides. The deduced amino acid sequence of the edited IKU1 gene showed the introduction of stop codon responsible for the premature termination of the protein transduction. So far 8 citrange plantlets have been recovered and confirmed as edited. These transformed plantlets are now in the juvenile phase and further analysis of their flowers and fruits features would confirm the role of IKU1 gene in IKU pathway and its potential for the obtainment of seedless cultivars. | |
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