S-Space College of Agriculture and Life Sciences (농업생명과학대학) Dept. of Food and Animal Biotechnology (식품·동물생명공학부) Theses (Master's Degree_식품·동물생명공학부)
Detection of CP4 5-enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) in soybean food products by affinity chromatography
대두 가공 식품으로부터 친화 크로마토그래피를 이용한 CP4 5-enolpyruvylshikimate-3-phosphate의 검출
- Issue Date
- 서울대학교 대학원
- 친화 크로마토그래피; Cp4 5-enolpyruvylshikimate-3-phosphate synthase; 유전자 재조합 식품; Affinity chromatography; Shikimate 3-phosphate; Genetically modified organism
- Thesis (master`s)--서울대학교 대학원 :농생명공학부,2003.
- The Round-up Ready£¿ soybean, which has the tolerance against an herbicide,
glyphosate, is widely used for soybean processing. Recently, detection of
genetically modified (GM) food becomes more important because of mandatory
labeling. In general, there are two quantitation methods of genetically modified
soybean in soybean food products; one is based on PCR analysis at DNA level and
the other is an immunoassay for the CP4 5-enolpyruvylshikimate-3-phosphate
synthase (CP4 EPSPS) at protein level. However, these two methods have
limitations when GM soybean is processed to Korean traditional foods.
Therefore, development of a new detection method is needed for soybean food
products. Affinity chromatography could be a possibility as an alternative
detection method at protein level using the shikimate 3-phosphate as a ligand.
Especially, because CP4 EPSPS is intact after the heat-treatment for 90 min at
95 ¡ÆC, tofu can be applied onto affinity chromatography. An affinity
chromatography column was made by immobilizing shikimate 3-phosphate to the
beaded agarose gel through the carbodiimide condensation. Since the shikimate 3-
phosphate was not available commercially, it was synthesized enzymatically by
shikimate kinase II and purified by ion exchange chromatography. The protein
samples isolated from GM soybean were applied to an affinity chromatography
column and CP4 EPSPS was analyzed by SDS-PAGE. It seemed that CP4 EPSPS was
adsorbed onto affinity chromatography column. However, Because shikimate 3-
phosphate was purified with low level, the efficiency of affinity chromatography
column was not good. To prepare a large quantity of shikimate 3-phoshate, an
affinity chromatography can be a successful detection method for CP4 EPSPS from
GM soybeans and its food products. For the better analysis of CP4 EPSPS, the
changes of ligand immobilizing method and protein elution condition should be
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