Drought responsive genes in tea cultivars grown in Kenya

Abstract

Tea is one of the most popular non-alcoholic beverages worlwide, and a leading foreign exchange earner and source of livelihood to over three million people in Kenya. However, tea growing areas in Kenya experience abiotic stresses with drought been the most predominant. Tea plants tolerate drought through poorly understood physiological, cellular/biochemical and molecular processes. Development of tea cultivars adapted to water-deficit stress greatly relies on an understanding of mechanisms of plant responses. Use of sequencing technologies provide a fast, cost effective, and reliable approach to generate large expression datasets for functional genomic analysis in plants exposed to drought. In the present study, 18-months old seedling tea from eight cultivars, tolerant and susceptible to drought, were subjected to three levels of treatment consisting of high (34%), moderate (26%) and low (18%) soil moisture content in a rain-out shelter. The experiment was designed in a complete randomized block design with three replications. After three months of exposure to treatment, physiological parameters (leaf water status, shoot growth and gas exchange parameters), biochemical parameters (proline and glycinebetaine levels) were determined. The data generated were subjected to two-way anova using Genstat. For molecular analysis, total RNA extracted was from tolerant and susceptible cultivars under stressed and unstressed conditions. The extracts were used to isolate mRNA which was reverse transcribed to complementary DNA. The sequences/reads generated from the cDNA libraries using 454 GLX sequencer were analysed in silico using bioinformatics tool. There was a reduction in shoot growth, leaf relative water content, shoot water potential and gas exchange parameters that varied significantly, P<0.05, amongst the cultivar and treatment in all the cultivars under study. However, proline and glycinebetaine increased with stress. Cultivar TRFK 306 accumulated the highest, 0.53μmol/g FW, level at low (18%) SMC whereas TN14-3 accumulated the least, <0.1 μmol/g FW, amounts. Glycinebetaine accumulation was high, 2.4mMoles/g DW, in AHP S15/10 and lowest (1.4 mMoles/g DW) in TRFK 301/5 under low soil moisture content. Accumulation of the two osmolytes also varied significantly (P<0.05) amongst treatment with the lowest soil moisture content inducing higher accumulation of the two biochemicals analysed. The 232,853 high quality raw reads generated from the sequencer were quality-filtered, trimmed and assembled into 460 long transcripts (contigs). The contigs were annotated by BLAST similarity search against proteins in Arabidopsis proteome. Based on Gene ontology analysis, drought response related transcripts including Heat shock proteins, Hsp70, antioxidant molecules such as Superoxide dismutase, catalase and peroxidise, signal tranducers, Calmoduline like protein, and vii Galactinol synthase (Gols4) were induced in the water stressed plants. In conclusion, molecular changes in tea plants bring about the physiological and biochemical changes. The physiological and biochemical changes observed thus were a clear indicator that water stress response is controlled at the genetic level.