A study on the extraction procedure and electrophoresis via SDS-page of the total proteins from leaves of some orchids from the Philippines

Date

6-1999

Degree

Bachelor of Science in Biology

Major Course

Major in Cell Biology

College

College of Arts and Sciences (CAS)

Adviser/Committee Chair

Carmina C. Manuel

Abstract

This study was done to evaluate and compare protein extraction methods (with and without protein precipitation using acetone, as well as with and without centrifugation of the sample) and three homogenizing buffer systems (0.2 M phosphate buffer, p11 7.0, 0.1 M tris-HCI buffer, pH 6.8 and phosphate buffered saline) for the electrophoresis via SOS-PAGE of total proteins from the leaves of orchids collected from the Philippines. The effect of the protein extraction methods and buffer systems were compared in terms of the amount of protein extracted (pg/a1) and the resolution and number of protein bands observed among 30 orchid plants consisting of 16 species. Protein precipitation with the use of acetone caused a significant decrease in the amount of proteins obtained among the sampled orchids. Centrifugation likewise decreased the amount of proteins obtained as well as the number of bands resolved but not significantly except for the significant decrease in the number of bands for samples homogenized in tris-HCI buffer and then centrifuged. The comparison of phosphate buffer, tris-HCI buffer and phosphate buffered saline for uncentrifuged samples showed non-significant difference in terms of the protein concentration but exhibited significant differences in the mean number of protein bands resolved, being highest in tris-HCI and lowest in phosphate buffered saline. On the other hand, for the centrifuged samples homogenized in phosphate and tris-HCl buffers, there were no significant differences observed in terms of the protein concentration and number of bands obtained. It is interesting to note, however, that in all of the 30 samples used for comparison between tris-HCI buffer and phosphate buffer, similarity indices (SI) fall below 50%. The average SI of the orchid plants extracted with tris-HCI buffer was at 24.18% while those extracted with phosphate buffer was at 26.95%. SI within species were also below SO% except for Bulbophyllum sp. extracted in phosphate buffer (75%). The low SI values obtained suggest a high genetic variation among the orchids examined, but this may also be due to the small number of replicates considered for each species.

Language

English

Location

UPLB Main Library Special Collections Section (USCS)

Call Number

Thesis

Document Type

Thesis

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