Estimation of endogenous indole-3-acetic acid in leaf explants and a protocol for direct regeneration in Lilium cv. Pavia

Abstract

A method for regeneration via direct organogenesis in Lilium cv. Pavia using leaf primary explants has been established. Outer leaves from in vitro plantlets of 10 to 12 cm were used as the primary explant for direct organogenesis. Shoots were directly regenerated from leaf explants within 25 d of culture on Murashige and Skoog (MS) augmented with 4.5 μM 2, 4-dichlorophenoxyacetic acid (2, 4-D), 5.4 μM α-naphthaleneacetic acid (NAA), and 2.2 μM 6-benzylaminopurine (BAP). Different regions of the same leaf explant exhibited varied responses. The acropetal 3 to 4 cm of the leaf (L1) produced the maximum number of shoots (75%) indicating its high regeneration ability whereas the response of middle portion (L2) and basipetal portion of the leaf (L3) exhibited a significantly decreasing trend of shoot induction with 37.5 and 12.5%, respectively. HPLC analysis of endogenous indole-3-acetic acid (IAA) concentration established significantly higher concentration in the acropetal segments (L1). The in vitro regenerated plants were rooted in half strength MS media and were successfully hardened and field transferred. Our study established that different regions of the same leaf produce significantly different responses for shoot induction and this is attributed to the variable levels of endogenous IAA. This quick method of direct organogenesis can be applied for commercial multiplication of this important hybrid.