Evaluation of 21 different media on shoot regeneration in 11 cultivars of citrus using juvenile tissue

Authors: MEIRA, FILIPE - Oak Ridge Institute For Science And Education (ORISE); SHAO, MIN - Oak Ridge Institute For Science And Education (ORISE); Niedz, Randall; Thomson, James - Jim

Submitted to: Plant Cell Tissue and Organ Culture
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/16/2024
Publication Date: 6/24/2024
Citation: Meira, F.S., Shao, M., Niedz, R.P., Thomson, J.G. 2024. Evaluation of 21 different media on shoot regeneration in 11 cultivars of citrus using juvenile tissue. Plant Cell Tissue and Organ Culture. 158. Article 8. https://doi.org/10.1007/s11240-024-02785-7.
DOI: https://doi.org/10.1007/s11240-024-02785-7

Interpretive Summary: Tissue transformation is a process that allows DNA modification of the plant genome. The modification can be used for a host of improvements such as, but not limited to, fruit color change and increased disease resistance. With the threat of HLB to commercial citrus production in California, improved disease resistance is the main drive for this project. Transformation technology in citrus is difficult, time consuming and limited to a few non-commercially relevant cultivars. Without improvements to the citrus transformation process, when a cure for huanglongbing (HLB) is discovered that requires genome modification, a majority of the existing citrus cultivars will lose commercial viability due to their inability to be modified. The project examined 21 unique media recipes to enhance tissue regeneration in 11 citrus cultivars.

Technical Abstract: Citrus is a globally cultivated fruit crop with oranges accounting for over half of the Citrus production. However, Huanglongbing (HLB), a devastating disease, is seriously affecting the citriculture industry. Agrobacterium-mediated transformation of juvenile materials is an option to combating this disease and has been a successful way of engineering Citrus. However, the regeneration of shoots remains a bottleneck in the application to commercial Citrus varieties. Citrus in vitro organogenesis is influenced by various internal and external factors, making it challenging to standardize the protocol. The genotype and season dependency of the explant make it challenging to standardize a protocol. The regeneration potential of the explant is specifically correlated with the physiological conditions of the donor plant. The success of recovering transgenic Citrus plants from transformed cells is increased by use of a culture medium an adequate mineral supply and combination of plant growth regulators. This study aimed to optimize the in vitro organogenesis protocol for shoot regeneration from the juvenile tissue of Citrus. Twenty-one distinct compositions in 11 Citrus cultivars were tested to determine the optimal media formulation for shoot regeneration. Regeneration of shoots statistical analyses were performed on the 21 media (1st round) and 11 media (2nd round) experiments utilizing 11 cultivars. The number of shoots from juvenile tissue was used in the third-month to evaluate cultivar efficiency. The results of the study showed that the success of shoot regeneration from Citrus juvenile tissue is highly dependent on the composition of the culture medium and cultivar. The optimal media formulation resulted in a high number of shoots from juvenile tissue in all 11 cultivars tested, indicating that it could be used for an enhanced in vitro organogenesis protocol for Citrus. In conclusion, the optimization of the in vitro organogenesis protocol for shoot regeneration from Citrus juvenile tissue is offered for improved rates of regeneration and the genetic modification necessary to develop HLB resistant Citrus types.