Genotype to Chemotype Analysis in Sage (Salvia officinalis L., Lamiaceae)

Chemotypes (or chemovars) are chemically distinct entities within a plant species based on genetic variation. Their occurrence is in mono- and sesquiterpenes (principal constituents of essential oils) exceptionally frequent. In almost all chemotypes the genetic background of their formation is still unknown. Salvia officinalis L. (garden sage, Dalmatian sage, Lamiaceae), a medicinal and aromatic plant whose numerous different bioactivities are partly based on its essential oil, is a prime example of the occurrence of a multitude of chemotypes. The genetic resources already available for Dalmatian sage (gene sequences and protein structures of monoterpene synthases, gene regulation, etc.) permit here an in depth elucidation of the genetic background of chemotype formation. A population of garden sage from Central Albania will be the model population in focus to study this background because this population is polymorphic in a specific sensorially important essential oil composition. In the beginning, the natural variation of the target genes (different monoterpene synthases) on the genomic level will be compiled including their 5Ą¯ and 3Ą¯ UTRs and the promoter regions. The major difficulty is the occurrence of multiple copies of each gene in the genome whose exact number is still unknown. Therefore, a complex qPCR and sequencing approach is foreseen to detect haplotypes within individuals. After an inventory of haplotypes, an extensive analysis of the transcriptome will follow in order to find out which haplotypes are expressed in different chemotypes and plant tissues. In a next step, the association between expressed haplotypes and chemotypes will be studied first in silico and then in an in planta transient expression system. Furthermore, promoter regions will be analyzed, because differences here could also be responsible for the chemical polymorphisms. Deletion analysis of the promoter regions shall further clarify, which minimal requirements are necessary for gene expression of monoterpene synthases. Two bioactive monoterpenes in sage, alpha and beta-thujone, have potential neurotoxic effects when consumed in excess. Therefore, the thujone chemotypes will be studied in a neuronal cell culture assay to determine if the chemotypes show different activity in this assay and if the plant extract has the same activity as pure compounds. In the end our results will not only explain the correlation between genotype and a specific chemotype in Dalmatian sage but shall also enable a faster and straightforward exploration of further genotype\chemotype correlations in Dalmatian sage and many others of the numerous essential oil delivering plant species from the same family because in many of these closely related species, the basic genetic principle of chemotype could be similar. The known correlation between genotype and chemotype may also be important for quality assurance systems in pharmacy and the food sector, because this work could pave the way towards enables molecular markers to determine a specific chemotype without chemical analysis.