کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10843934 | 1069297 | 2005 | 8 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Refolding and purification of non-fusion HPT protein expressed in Escherichia coli as inclusion bodies
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موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
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چکیده انگلیسی
The gene encoding hygromycin B phosphotransferase (hpt) is a widely used selectable marker in the production of genetically engineered crops. To facilitate the safety assessment of this protein, the non-fusion hpt expression plasmid was constructed and introduced into Escherichia coli to produce enough quantity of the HPT protein. High level expressed HPT was achieved but most of the expressed protein aggregated as inclusion bodies. The inclusion bodies were washed, separated from the cells, and solubilized by 0.3% Sarkosyl. The protein was renatured by dilution and dialysis, and then purified by anion-exchange chromatography. The activity is 8Â U/mg protein and the purity is about 95%. Further studies showed that the microbially produced HPT protein had comparable molecular weight, immuno-reactivities, N-terminal amino acid sequences, and biological activities with those of the HPT produced by transgenic rice harboring hpt gene. All these results demonstrated the validity of utilizing the microbially produced HPT to assess the safety of the HPT protein produced in genetically engineered rice.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 41, Issue 1, May 2005, Pages 53-60
Journal: Protein Expression and Purification - Volume 41, Issue 1, May 2005, Pages 53-60
نویسندگان
Qin Zhuo, Jian-hua Piao, Rui Wang, Xiao-guang Yang,