Selection of High Phenolics-Containing Clones of Thyme (<i>Thymus vulgaris</i>L.) Using<i>Pseudomonas Sp</i>.

Abstract

A high phenolics-containing clonal line (T-12) of thyme (Thymus vulgaris L.) was isolated from a heterogeneous seed population by plant tissue culture techniques. This clonal line was isolated from among 10 clonal lines, with each originating from different genetically heterozygous, single-germinating seedlings. All clonal lines were generated via shoot organogenesis through adventitious bud proliferation from apex explants. Optimum shoot organogenesis was induced on Murashige and Skoog (MS) medium with benzyladenine (1 mg/L) as the growth hormone. Multiple shoots originating from apex explants of single heterozygous seedlings were further multiplied on the aforementioned benzyladenine-containing MS medium to subsequently generate a larger number of clonally identical plants. Shoots from each individual clonal line were inoculated with a novel Pseudomonas sp. Following growth on hormone-free MS medium for 25 days, total phenolics were determined spectrophotometrically. Using this approach, high phenolics-stimulated clonal line T-12 and moderate phenolics-stimulated clonal time T-16G were isolated. These clonal lines attained the higher level of phenolics following Pseudomonas inoculation and also had uninhibited shoot growth compared with the corresponding uninoculated control. Several low phenolics clonal lines, which had inhibited shoot growth in response to Pseudomonas sp., were also isolated. Thymol levels of uninoculated shoots of all clonal lines after 60 days of growth were also measured by gas chromatography−mass spectroscopy. The high-to-medium phenolics-containing clonal lines (T-12 and T-16G) had basal thymol levels in the range of 150 μg/g fresh weight (FW). The thymol levels of low phenolics-containing clonal lines were in the range 10−70 μg/g FW. This Pseudomonas sp.-mediated selection provides a potentially novel biotechnology based strategy to isolate high phenolics and thymol-containing clonal lines of thyme from a genetically heterogeneous population. Keywords: Biotechnology; clonal lines; micropropagation; phenolics; Pseudomonas sp.; thymol; Thymus vulgaris