In vitro screening of selected elite clones of Eucalyptus tereticornis Sm. for drought stress

Abstract

In the present study, in vitro screening of the four selected elite clones (‘KE8’, ‘CE2’, ‘T1’ and ‘Y8’) of Eucalyptus tereticornis for water stress (drought) tolerance was investigated. The proliferating multiple shoots were cultured on basal MS medium supplemented with 2.5 µM Benzyl adenine , 0.5 µM napthaleneacetic acid and different concentrations of mannitol (0-1000 mM). The shoot culture index of these clones was calculated under different level of stress and it was observed that shoot culture growth index of clone ‘KE8’ (141.42 %) was significantly higher than clone ‘Y8’ (106.07 %) on medium containing 1000 mM mannitol. The tolerant clone ‘KE8’ exhibited 72.79 % relative water content on medium containing 1000 mM mannitol whereas it was reduced to 60.29 % in sensitive clone ‘Y8’ on the same medium after a culture period of 28 days. It was also recorded that the sap of drought tolerant clone ‘KE8’ had a osmotic potential of -3.30 MPa, whereas sensitive clone ‘Y8’ had a -3.79 MPa on medium containing 1000 mM mannitol which had water potential of -3.16 MPa. In clone ‘KE8’, total chlorophyll and carotenoid contents of 116.88 µg g-1 fresh weight (FW) and carotenoid content of 15.671 µg g-1 FW were recorded on medium containing 1000 mM mannitol. However, significantly lower values for total chlorophyll (58.29 µg g-1 FW) and carotenoid contents (10.17 µg g-1 FW) were recorded for sensitive clone ‘Y8’ on the same medium and after the same culture period. Maximum proline accumulation (4 µmol g-1 FW) was observed for tolerant clone ‘KE8’ on medium containing 750 mM mannitol whereas, on the same medium and after same culture period, clone ‘Y8’ accumulated 3.2 µmol g-1 FW of proline . Maximum reducing sugars content (130.32 mg g-1 DW) was recorded in the cultures of clone ‘KE8’ on medium containing 1000 mM mannitol and on same medium drought sensitive clone ‘Y8’ accumulated 79.09 mg g-1 DW of reducing sugar. Similarly, total soluble sugar content also increased significantly with increasing concentration of mannitol, increase was more pronounced in tolerant clone than sensitive clone ‘Y8’. Increased antioxidant enzyme activity played a significant role in imparting drought stress tolerance in various clones of E. tereticornis. In this context, maximum superoxide dismutase (SOD) activity (11.09 U mg-1 protein ) was recorded in the cultures of clone ‘KE8’ on medium containing 1000 mM mannitol, whereas significantly lower SOD activity (6.99 U mg-1 protein) was recorded in the cultures of clone ‘Y8’. Further, the activities of other antioxidant enzymes such as catalase, peroxidase and ascorbate peroxidase were also studied and it was observed that the activities increased significantly with increasing concentration of mannitol. The sensitive clone ‘Y8’ also recorded higher levels of malondialdehyde content (21.06 µmol g-1 FW) on medium containing 1000 mM mannitol, indicating the extent of lipid peroxidation of cellular membrane due to the formation of reactive oxygen species. The present study explored the effect of mannitol on selected elite clones of Eucalyptus tereticornis for drought tolerance and studied various physiological and biochemical parameters to associate their role with drought tolerance. Taken together, the study resulted in the identification of tolerant clone ‘KE8’ which can be easily grown in areas with limited rainfalls