The formation of adventitious buds in vitro on micro-cross sections of hybrid Populus leaf midveins

Abstract

Micro-cross sections (MCS) of midveins from hybrid Populus leaves were1used to study adventitious bud formation in vitro. MCS of 100 μm, 200 μm, 300 μm, 400 μm, and 500 μm in length were made with a Lancer Vibratome and cultured in Woody Plant Medium (WPM). In the presence of naphthaleneacetic acid (NAA), 400 and 500-μm sections had equal bud forming capacity and produced as many adventitious buds (average four shoots/MCS) as a 1-cm explant under the same culture conditions. This means that MCS of midveins are approximately 25 times more efficient in shoot regeneration than a 1-cm explant. The conditions used were suboptimal for the 200-μm and 100-μm sections. Shoot numbers from the 200-μm MCS were considerably reduced and the 100-μm MCS did not form buds at all. The size of lamina remnants was critical with 1–2 mm on each side of the midvein being optimal for bud formation. The MCS (300 and 400 μm) were not only an effective system for production of adventitious shoots, but also allowed direct microscopic observations of early morphogenetic events in vitro. These direct observations indicate that the callus and buds are initiated from a group of small chlorophyllous cells located in areas of the midvein that are adjacent to the mesophyll cells.