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Proceedings Paper

Spectroscopic detection of fluorescent protein marker gene activity in genetically modified plants
Author(s): O. W. Liew; Jenny P. C. Chong; Anand K. Asundi
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Paper Abstract

This work focuses on developing a portable fibre optic fluorescence analyser for rapid identification of genetically modified plants tagged with a fluorescent marker gene. Independent transgenic tobacco plant lines expressing the enhanced green fluorescence protein (EGFP) gene were regenerated following Agrobacterium-mediated gene transfer. Molecular characterisation of these plant lines was carried out at the DNA level by PCR screening to confirm their transgenic status. Conventional transgene expression analysis was then carried out at the RNA level by RT-PCR and at the protein level by Western blotting using anti-GFP rabbit antiserum. The amount of plant-expressed EGFP on a Western blot was quantified against known amounts of purified EGFP by scanning densitometry. The expression level of EGFP in transformed plants was found to range from 0.1 - 0.6% of total extractable protein. A comparison between conventional western analysis of transformants and direct spectroscopic quantification using the fibre optic fluorescence analyser was made. The results showed that spectroscopic measurements of fluorescence emission from strong EGFP expressors correlated positively with Western blot data. However, the fluorescence analyser was also able to identify weakly expressing plant transformants below the detection limit of colorimetric Western blotting.

Paper Details

Date Published: 12 April 2005
PDF: 7 pages
Proc. SPIE 5852, Third International Conference on Experimental Mechanics and Third Conference of the Asian Committee on Experimental Mechanics, (12 April 2005); doi: 10.1117/12.621891
Show Author Affiliations
O. W. Liew, Singapore Polytechnic (Singapore)
Jenny P. C. Chong, Singapore Polytechnic (Singapore)
Anand K. Asundi, Nanyang Technological Univ. (Singapore)

Published in SPIE Proceedings Vol. 5852:
Third International Conference on Experimental Mechanics and Third Conference of the Asian Committee on Experimental Mechanics
Chenggen Quan; Fook Siong Chau; Anand Asundi; Brian Stephen Wong; Chwee Teck Lim, Editor(s)

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