Rapid Microwave Processing of Winter Cereals for Histology Allows Identifi cation of Separate Zones of Freezing Injury in the Crown

In histological studies, microwave processing of tissue considerably shortens the time required to prepare samples for observation under light and electron microscopy. However, plant tissues from different species and different regions of the plant respond differently to microwave processing, making it impossible to use a single protocol for all plant tissue. The crown of winter cereals such as rye (Secale cereale L.), wheat (Triticum aestivum L.), barley (Hordeum vulgare L.), and oats (Avena sativa L.) is the below-ground portion of the stem that overwinters. It is composed of numerous types of cells with an organizational pattern that is similar to other grasses. When we used microwave protocols that were developed for other plant tissues, winter cereal crown tissue shattered and crumbled when sectioned. This study reports a procedure developed to process winter cereal crowns for histological observations. Using this microwave protocol, samples were prepared in 1 d as compared to 2 wk using traditional protocols. This enabled many more samples to be processed and allowed us to identify four overlapping zones of response to freezing within the crown. Results of varying time, temperature, and microwave wattage during fi xing, dehydrating, and embedding in paraffi n are described. High quality sections from the crowns of oat, barley, wheat, and rye indicate that this procedure is valid for all winter cereals. Since crown tissue is similar across all grass species, we predict that the protocol will be useful for other grasses as well. D.P. Livingston III, USDA and North Carolina State University, 840 Method Rd, Unit 3, Raleigh, NC 27695; T.D. Tuong, USDA and North Carolina State University, 3127 Ligon Street, Raleigh, NC 27607-7631; C.H. Haigler, Dep. of Crop Science and Dep. of Plant Biology, North Carolina State University, Campus Box 7620, Raleigh, NC 27695; U. Avci, Dep. of Crop Science, Campus Box 7620, North Carolina State University, Raleigh, NC 27695, current address: University of Georgia, 2048 CCRC, Athens, GA 30602; S.P. Tallury, North Carolina State University, 840 Method Rd, Unit 3, Raleigh, NC 27695. Received 27 May 2009. *Corresponding Author ([email protected]). Abbreviations: Ethanol, EtOH; recombinant inbred line, RIL; tertiary butyl alcohol, TBA. Published in Crop Sci. 49:1837–1842 (2009). doi: 10.2135/cropsci2009.02.0077 © Crop Science Society of America 677 S. Segoe Rd., Madison, WI 53711 USA All rights reserved. No part of this periodical may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher. Permission for printing and for reprinting the material contained herein has been obtained by the publisher.