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KMID : 0545120070170040681
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Journal of Microbiology and Biotechnology
2007 Volume.17 No. 4 p.681 ~ p.684
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Quantitative Analysis of Phosphinothricin-N-acetyltransferase in Genetically Modified Herbicide Tolerant Pepper by an Enzyme-Linked Immunosorbent Assay
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Shim Youn-Young
Shin Weon-Sun
Moon Gi-Seong
Kim Kyung-Hwan
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Abstract
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An immunoassay method was developed to quantitatively detect phosphinothricin-N-acetyltransferase (PAT) encoded by the Bialaphos resistance (bar) gene in genetically modified (GM) pepper. The histidine-tagged PAT was overexpressed in Escherichia coli M15 (pQE31-bar) and efficiently purified by Ni2+ affinity chromatography. A developed sandwich enzyme-linked immunosorbent assay (S-ELISA) method (detection limit: 0.01 μg/ml) was 100-fold more sensitive than a competitive indirect ELISA (CI-ELISA) method or Western blot analysis in detecting the recombinant PAT. In real sample tests, PAT in genetically modified herbicidetolerant (GMHT) peppers was successfully quantified [4.9± 0.4 μg/g of sample (n=6)] by the S-ELISA method. The SELISA method developed here could be applied to other GMHT crops and vegetables producing PAT.
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KEYWORD
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Phosphinothricin-N-acetyltransferase, bar gene, GM pepper, sandwich ELISA
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