Influence of polyamines on hyperhydricity reversion and its associated mechanism during micropropagation of China pink (
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RESEARCH ARTICLE
Influence of polyamines on hyperhydricity reversion and its associated mechanism during micropropagation of China pink (Dianthus chinensis L.) R. Sreelekshmi1 • E. A. Siril1
Received: 28 April 2020 / Revised: 28 July 2020 / Accepted: 17 September 2020 / Published online: 28 September 2020 Ó Prof. H.S. Srivastava Foundation for Science and Society 2020
Abstract Hyperhydricity (HH) is a physiological disorder that frequently occurs in plant tissue cultures, affecting healthy growth and development of clonal plants. The primary cultures raised in Murashige and Skoog (MS) medium supplemented with 2.5 lM N6-benzyladenine (BA) produced normal microshoot (6.3 shoots/ culture) with least HH. However, the third subculture onwards, HH becomes a major problem. The role of ethylene on HH induction through stomatal closure mechanism were proved by the supplementation of ethephon (5 lM) in the culture medium containing 2.5 lM BA. In the present study, the application of polyamines (putrescine, spermidine, or spermine) to minimize the HH was examined. Supplementation of 5 lM spermine in MS medium significantly reduced the percentage of HH to 0.33%, in contrast to control (100%), while a maximum number of healthy reverted shoots (11.0) were observed in 5 lM spermidine treatment. The addition of polyamines effectively reduced H2O2 content (50%) characterized by increased chlorophyll content with proper stomatal morphology. The relative gene expression profile of ethylene biosynthesis genes, 1-Aminocyclopropane-1-carboxylase synthase (ACS1) and 1-Aminocyclopropane-1-carboxylic acid oxidase (ACO1) at 5 lM spermine added medium was 1.09 and 1.3 over normal (1) or HH cultures (1.93 and 2.53) respectively, and thus directed restoration of normal morphology of shoots. The present finding in brief, forward a novel method to regulate HH in terms of endogenous ethylene by adopting polyamines exposure and the & E. A. Siril [email protected] 1
Department of Botany, University of Kerala, Kariavattom, Thiruvananthapuram 695581, India
procedure can be applied to many other plants facing similar HH problems. Keywords ACC synthase Ethephon Spermine Stomatal bioassay Vitrification
Introduction Plant tissue culture is a technique widely employed in the field of agriculture, horticulture, plant breeding, and forestry due to its accepted advantages over conventional tools. The in vitro propagation systems can be of beneficial use in many plant species for rapid multiplication, germplasm conservation, and genetic manipulation. However, the in vitro propagation can sometimes be problematic due to the development of various tissue abnormalities. Hyperhydricity (HH) or vitrification is one of the significant problems that impose extreme oxidative stress to in vitro growing tissues (Saher et al. 2005). The hyperhydric (glassiness) plantlets are easily breakable, and shoots are translucent, thick, and pale green. These physiological disorders are characterized under extreme culture conditions viz., high relative humidity, weak g
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