Induced polyploidy and broad variation in phytochemical traits and altered gene expression in Salvia multicaulis

Induced polyploidy is nowadays an important strategy in plant breeding and for the development of new crops. Salvia multicaulis Vahl is a valuable medicinal plant that produce precious bioactive metabolites including triterpenic acids (TAs) and phenolic compounds. Hence, at first, for selecting elite lines, both HPLC and GC-MC analyses were performed on fourteen S. multicaulis lines. Then, seeds of selected lines of S. multicaulis were exposed to different concentrations (0.00, 0.05, 0.1, and 0.2 %) of colchicine for 24 or 48 h. The flow cytometric analysis and chromosome counting were used to confirm ploidy level of tetraploid control (2n = 4x = 28, 2C DNA = 1.36 pg) and hexaploid (2n = 6x = 42, 2C DNA = 1.97 pg) plants after seven-month. For the first time, the effects of in vitro polyploidization on morphological characteristics, TAs and phenolic acid contents as well as on the expression of six TAs biosynthesis related genes were investigated. The highest efficiency of hexaploidy (12.76 %) was achieved 48 h after exposure to 0.1 % colchicine concentration. The hexaploid plants showed different growth traits compared with those of tetraploid control plant; indeed, hexaploid plants had leaves with a darker green color, a lower trichome density, and lower plant height and root length. Moreover, there was a significant increase in rosmarinic acid and caffeic acid content in hexaploid plants compared with tetraploid control plants. Also, the increase of oleanolic acid (1.33 fold) content in hexaploids was associated with a significant increased expression of squalene synthase (SQS) and β-amyrin synthase (BAS) genes in hexaploid plants. Nevertheless, a significant decreased expression of squalene epoxidase (SQE), mixed-function amyrin synthase (MFAS), and lupeol synthase (LUS) was observed in hexaploid plants, that led to a reduced content of ursolic acid and betulinic acid compared with tetraploid control plants. These results confirmed that polyploidization is a breeding method with stochastic results in secondary metabolites production and gene expression related to biosynthetic pathways.