Factors influencing the survival of developing embryos of theobroma cacao L. (Malvaceae) in cryogenic storage

Authors: Crane, Jennifer; Irish, Brian; Walters, Christina

Submitted to: International Society for Horticultural Science Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 8/13/2013
Publication Date: 8/13/2013
Citation: Crane, J., Irish, B.M., Walters, C.T. 2013. Factors influencing the survival of developing embryos of theobroma cacao L. (Malvaceae) in cryogenic storage. International Society for Horticultural Science Meeting. p. 72.

Interpretive Summary:

Technical Abstract: Cacao, Theobroma cacao L., is native to tropical South American rainforests and is the source of chocolate. Ex situ conservation of this economically important species and its relatives is imperative to prevent genetic erosion resulting from diminished suitable habitat and increased pressure from a wide range insect and disease pests. Collections of elite cacao germplasm are maintained in large field plots and research is underway to preserve these specific genotypes using somatic embryos (e.g., Adu-Gyamfi and Wetten, 2012, CryoLetters 33: 494) or vegetative cuttings (Gayle Volk, personal communication). However, conservation of the full range of genetic diversity is best approached by preserving sexually-derived germplasm. Impeding this conservation effort for cacao is the recalcitrant nature of the seeds, which are quickly killed in conventional freezers (-18oC) used by most genebanks. In addition to the propensity of cacao storage lipids to crystallize, the desiccation sensitivity of embryo cells necessitates cryogenic storage strategies which optimize embryo size, moisture content, cooling rate to liquid nitrogen temperature and cryoprotectant treatments. Zygotic embryos develop over an approximately six month period and mature embryonic axes may accumulate as much as 12 mg dry mass. The combination of large sample mass and high water content at maturity (about 2 g H2O/ g dry mass) make it unfeasible to cryopreserve intact fully mature embryonic axes. Less mature axes, collected approximately three months after flowering, are smaller, but contain considerably more water (6-9 g H2O/g dry mass) that must be removed using pre-culture or cryoprotectants. These small immature axes may also be difficult to grow in culture. Differences in embryogenic patterns among genetic lines and harvest seasons make it difficult to precisely target pods at the optimum developmental stage. The current work seeks to describe the effect of interacting factors, such as water relations, cellular constituents and cell structure on survival of embryonic axes of cacao that are cryoexposed at different maturity stages. Future research will focus on comparing these factors, as well as cryoprotectant concentration and exposure times, in zygotic and somatic embryos to determine the conditions that limit successful cryopreservation.