Development of an efficient tissue culture protocol for callus formation and plant regeneration of wetland species Juncus effusus L.

Abstract

Wetland species mat rush (Juncus effusus L.) is an important economic plant, but no information is available regarding plant regeneration, callus induction, and its proliferation from in vitro seed grown plantlets. The present study investigates the effects of growth regulator combinations and medium innovation on tissue culture system of five mat rush varieties. Addition of N6-benzyladenine (BA) and 2,4-dichlorophenoxyacetic acid (2,4-D) in Murashige and Skoog (MS) medium showed significantly positive effect on callus proliferation, plant regeneration, and its multiplication compared to the medium devoid of BA. The highest callus induction frequency (80.95%, 90.48%, 75.40%, 70.83%, and 83.33%) was observed in MS medium containing 0.5 mg L −1 (2.2 μM) BA in Yinlin-1, Nonglin-4, Gangshan, Taicao, and Taiwan green, respectively. Various growth regulator combinations with successive subculture (medium replacement) were found essential to develop organogenic calluses and to regenerate shoots. The combination of 0.1 mg L −1 BA (0.4 μM) and 2 mg L −1 2,4-D (9.0 μM) in MS medium was found best for callus proliferation for all the varieties under trial. The plant regeneration required two steps involving successive medium replacements as well as optimal hormonal balances. Successful plant regeneration (over 70%) was observed only by transferring the organogenic callus from regeneration medium I [MS medium containing 0.5 mg L −1 BA (2. μM) and 1.0 mg L −1 kinetin (KT; 4.6 μM)] to the regeneration medium II [MS medium containing 0.5 mg L −1 BA (2.2 μM), 1.0 mg L −1 KT (4.6 μM) and 3.0 mg L −1 indoleacetic acid (IAA; 17.1 μM)]. Our results confirmed the importance of the ratio of auxin (IAA) to cytokinin (BA and KT) in the manipulation of shoot regeneration in J. effusus L. The maximum plant survival frequency and multiplication rates (90.97% and 5.40 and 94.23% and 8.25) were recorded in the presence of 0.5 mg L −1 BA (2.2 μM) in the 1/2 MS multiplication medium for the varieties of Nonglin-4 and Taicao, respectively. About 100% survival rate was also observed for all the varieties in soil conditions. The efficient plant regeneration system developed here will be helpful for rapid micropropagation and further genetic improvement in J. effusus L.

Effect of Plant Growth Regulators on Callus, Cell Suspension and Cell Line Selection for Flavonoid Production from Pegaga (<i>centella asiatica</i> L. urban)

“…Xu et al (2009) 2,4-D and 1.5 mg L −1 BAP successfully induced yellowish callus on the 28th day. Notably, there was a significant reduction rate in callus induction response when auxin was applied together with cytokinin.…”

Induction of tetraploidy in Juncus effusus by colchicine

“…Basal culm segments (approximately 0.5 -1.0 cm in length) were treated with various concentrations of colchicine (0, 50, 100 and 500 mg dm -3 ) for 6, 12 and 24 h. The treated segments were transferred to 1/2 strength Murashige and Skoog (1962; MS) medium with 0.5 mg dm -3 benzyladenine (BA). Eight segments per flask were grown under 16-h photoperiod, irradiance of 100 μmol m -2 s -1 , and temperature of 19 ± 2 °C for three weeks (for detail see Xu et al 2009). Then the basal segments of fully developed plantlets were transferred to fresh MS media.…”

Insights into cadmium induced physiological and ultra-structural disorders in Juncus effusus L. and its remediation through exogenous citric acid

“…Shen, University of Nottingham Ningbo, Ningbo, Zhejiang province, China. Until sowing in Zhejiang University, Hangzhou, China, seeds were stored in the dark at 4 • C. Seeds were surface sterilized according to Xu et al [21] using a solution containing 0.05 g of potassium permanganate (KMnO 4 ) in 80 mL sterile water for 72 h transferred to ethanol (70%) for another 60 min, and washed with sterile distilled water. The seeds were then shifted into 1.0% sodium hypochlorite (NaClO) solution, containing 2 drops of Tween-20 and kept shaking for 20 min prior to three washings with sterile distilled water.…”