Sreelekshmi R, Siril E A
Department of Botany, University of Kerala, Kariavattom, Thiruvananthapuram, 695581 India.
Physiol Mol Biol Plants. 2020 Oct;26(10):2035-2045. doi: 10.1007/s12298-020-00885-0. Epub 2020 Sep 28.
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 µM N-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 µM) in the culture medium containing 2.5 µM BA. In the present study, the application of polyamines (putrescine, spermidine, or spermine) to minimize the HH was examined. Supplementation of 5 µM 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 µM spermidine treatment. The addition of polyamines effectively reduced HO 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 µM 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 procedure can be applied to many other plants facing similar HH problems.
玻璃化现象（HH）是植物组织培养中经常出现的一种生理紊乱现象，影响克隆植物的健康生长和发育。在添加了2.5 μM N-苄基腺嘌呤（BA）的Murashige和Skoog（MS）培养基中培养的初代培养物产生了正常的微芽（每个培养物6.3个芽），玻璃化现象最少。然而，从第三次继代培养开始，玻璃化现象就成了一个主要问题。通过在含有2.5 μM BA的培养基中添加乙烯利（5 μM），证明了乙烯通过气孔关闭机制在诱导玻璃化现象中的作用。在本研究中，研究了应用多胺（腐胺、亚精胺或精胺）来减少玻璃化现象。在MS培养基中添加5 μM精胺可使玻璃化现象的百分比显著降低至0.33%，而对照组为100%；同时，在5 μM亚精胺处理中观察到的健康恢复芽的数量最多（11.0个）。多胺的添加有效地降低了过氧化氢含量（50%），其特征是叶绿素含量增加且气孔形态正常。在添加5 μM精胺的培养基中，乙烯生物合成基因1-氨基环丙烷-1-羧酸合酶（ACS1）和1-氨基环丙烷-1-羧酸氧化酶（ACO1）的相对基因表达谱分别为1.09和1.3，高于正常（1）或玻璃化培养物（1.93和2.53），从而促使芽的形态恢复正常。简而言之，本研究结果提出了一种通过多胺处理来调节内源性乙烯从而控制玻璃化现象的新方法，该方法可应用于许多面临类似玻璃化问题的其他植物。
