biologia plantarum

Biologia plantarum 53:360-364, 2009 | DOI: 10.1007/s10535-009-0067-2

In vitro organogenesis and plant regeneration from unpollinated ovary cultures of Azadirachta indica

P. Srivastava¹, M. Singh¹, P. Mathur^1,2, R. Chaturvedi^1,*

¹ Department of Biotechnology, Indian Institute of Technology, Assam, India

² School of Biotechnology, VIT University, Vellore, India

A novel method of organogenesis in neem (Azadirachta indica A. Juss.) from unfertilized ovaries is described. The Murashige and Skoog's (MS) medium with 9 % sucrose, 1 µM 2,4-dichlorophenoxyacetic acid (2,4-D) and 5 µM 6-benzylaminopurine (BAP) was the best for callus induction from unfertilized ovaries. However, further proliferation of callus occurred better on MS medium supplemented with 0.5 µM 2,4-D either alone or in combination with 4.5 µM kinetin. Maximum shoot regeneration (78 %) was observed when calli, induced from ovaries of 4 mm size flower buds and proliferating on MS + 0.5 µM 2,4-D, were subcultured to MS medium containing 5 µM BAP. Histological analysis revealed that 4 mm sized flower bud corresponds to a 2-nucleate stage of embryo sac. The shoots were then multiplied by forced axillary branching on MS medium supplemented with 1.0 µM BAP and 250 mg dm^-3 casein hydrolysate. The shoots could be rooted on 1/4 strength MS medium supplemented with 0.5 µM indole-3-butyric acid (IBA) at a frequency of 79 %. Cytological analysis by root tip squash preparations revealed that all the plantlets were diploids. These plants were subsequently hardened and established in soil with transplantation rate of 81.8 %.

Keywords: adventitious shoot proliferation; auxins; callus culture; casein hydrolysate; cytokinins; embryo sac; histology; ovary culture

Subjects: auxins; Azadirachta indica; cytokinins; ex vitro transfer; in vitro culture, callus; in vitro culture, indirect regeneration

Received: September 30, 2007; Accepted: March 25, 2008; Published: June 1, 2009Show citation