Production and purification of two recombinant proteins from transgenic corn

被引:77
作者
Kusnaki, AR
Hood, EE
Witcher, DR
Howard, JA
Nikolov, ZL
机构
[1] Iowa State Univ, Dept Food Sci & Human Nutr, Ames, IA 50011 USA
[2] Iowa State Univ, Dept Agr & Biosyst Engn, Ames, IA 50011 USA
[3] Iowa State Univ, Ctr Crops Utilizat Res, Ames, IA 50011 USA
[4] ProdiGene Inc, College Stn, TX 77845 USA
[5] Pioneer Hi Bred Int Inc, Johnston, IA 50131 USA
关键词
D O I
10.1021/bp970138u
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
This study reports the production, purification, and characterization of recombinant Escherichia coli beta-glucuronidase (GUS) and chicken egg-white avidin from transgenic corn seed. The avidin. and gus genes were stably integrated in the genome and expressed over seven generations. The accumulation levels of avidin and GUS in corn kernel were 5.7% and 0.7% of extractable protein, respectively. Within the kernel, avidin and GUS accumulation was mainly localized to the germ, indicating possible tissue preference of the ubiquitin promoter. The storage-stability studies demonstrated that processed transgenic seed containing GUS or avidin can be stored at 10 degrees C for at least 3 months and at 25 degrees C for up to 2 weeks without a significant loss of activity. The heat-stability experiments indicated that GUS and avidin in the whole kernels were stable at 50 degrees C for up to 1 week. The buffer composition also had an affect on the aqueous extraction of avidin and GUS from ground kernels. Avidin was purified in one step by using 2-iminobiotin agarose, whereas GUS was purified in four steps consisting of adsorption, ion-exchange, hydrophobic interaction, and size-exclusion chromatography. Biochemical properties of purified avidin and GUS were similar to those of the respective native proteins.
引用
收藏
页码:149 / 155
页数:7
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