Analysis of the cross section of the Thuja cone seed using light microscope (LM) and scanning electron microscope (SEM)

Introduction:

Thuja Linnaeus 1753, genus of coniferous trees and division of Pinophyta, are commonly known as cedar (Figure 1), and found in North America, East Asia and Europe. They are the biggest group in gymnosperms. These monoecious trees, also known as arborvitae meaning “tree of life” in Latin, grow in North America, East Asia and Europe. Trees can be up to 15 meters tall. The protruding resin glands from Thuja produce strong aromatic smell. The cones in yellowish green, sized of 1.5 cm, are located underneath the branch of the evergreen, scale-like overlapping leaves lying opposite in pairs from the branch. The seed cones start developing from mid April and become mature in October. The small coniferous (fruits) cones are important because they consist of reproductive winged seeds, which are released by opening of the cone scales when dried and brown (Figure ). The seeds are fertilized ripened ovule of gymnosperm, consisting of embryo, nutritive tissue and seed coat. Thuja spp. can be widely used for various therapeutic purposes and ornamental decorations.

Materials and Methods:

Thuja cone was obtained from the Mount Allison campus during maturation season, the beginning of October. The seed cone was cut (sliced in the middle) cross-section and examined under a light microscope (LM). The LM cross-section view of the cone and the seed are illustrated in Figure (3) and (4) respectively. The sample was then prepared for inspection using a scanning electron microscope (SEM). First, the sample was sliced into a small thin piece cross-section in the middle. Then, the sample was fixed by a cold (4° C) 2.5% glutaraldehyde fixative solution in 0.1M phosphate buffer, pH 6.8, for one hour. The fixed sample was rinsed in buffer (phosphate buffer?) for four times at a room temperature. The specimen was dehydrated in each 20, 50, 70, 85 and 95% ethanol for 10 minutes and then 4 times in 100% anhydrous ethanol for 10 minutes. After that, the sample was collected on a filter and the critical point dried with liquid carbon dioxide. The specimen was mounted on aluminum stubs using double-sided tape and carbon paint with approximately 20 nm, gold in Hummer 6.2 sputtering unit by using gas. The sample was examined using a JEOL JSM-5600 SEM operating at 10 kV and a working distance of 46 mm. The SEM micrograph of the cross-section of Thuja female cone was shown in Figure (5). Along with it, cross-section of cone seed and a pouch are shown in Figure (6) and (7) respectively.

Results:

The cross-section of the cone under the light microscope appears as Figure ( ). The tetra porous seeds are attached at the axis, yet they are separated by the cone scales. The yellowish green Thuja cone is woody, leathery, or fleshy scales until the mid October. Several resin duct pores and air pouches are observed over the cross-section plane of the fresh green cone by observing the oozing of the clear liquid on the surface.

However, by the beginning of November, hardly could the green cones be observed. A dry brown cone seed at the maturation appears as Figure ( ). According to the observation by using LM, the cone has 4 seeds. Each has two seed wings and is separated by cone scale (Figure 4). Nine cone scales are observed when dried.

SEM Figure (4) shows an uneven surface of the cone seed, which was be observed by a light microscope. Some of the several holes and hollow areas observed on the cross-section surface could be resin ducts (Figure 6) surrounded by living cells of epithelium and some air pockets (Figure 6).

Conclusion:

The SEM micrograph did not show all the possible cross-section of the Thuja cone. By using LM, the different images of slicing could be observed. The number of seeds that could be observed on cross-section plane depends on the distance of the plane from the center of the cone was cut. The seeds observed were much smaller when they are cut further away from the central on cross-section. In addition, depending on the axial or radial section, the image of the seeds and scales are different.

Collaboration of LM and SEM sample results give better understanding of the cross-section of the Thuja cone seed. LM gives the surface general morphological information, and the SEM micrographs provide more detailed three-dimensional images with higher resolution power.

Reference:

Answers.com as a source of encyclopedia info http://www.answers.com/topic/thuja

Earle, Christopher J. Thuja. (May 30, 2007). Retrieved September 7, 2007, from http://www.conifers.org/cu/th/index.htm

Powell G., Beardmore T. and Atlantic Forestry Centre Staff. New Brunswick Tree and Shrub Species of Concern: A Field Guide. Fredericton, N.B.: Atlantic Forestry Centre, c2002.

Raven Peter H., Evert Ray F. and Eichhorn Susan E. Biology of Plants (7^th edition). New York: Freeman W. H. and Company

Van De Graff Kent M., Rushforth Samuel R., and Crawley John L. A Photographic Atlas for the Botany Laboratory (4^th edition). Colorado 80110: Morton D.