Journal article
High-level expression of LMW-GS and α-gliadin genes promoted by the expressed tag sequence of 5′ end in Escherichia coli
Protein Expression and Purification, Vol.105, pp.54-60
2015
Abstract
Wheat storage protein genes, especially low molecular weight glutenin subunit (LMW-GS) and gliadin genes are difficult to be expressed in Escherichiacoli, mainly due to the presence of highly repetitive sequences. In order to establish a high efficiency expression system for these genes, five different expression plasmids combining with 9 genes, viz. 6 LMW-GS and 3 α-gliadin genes isolated from common wheat and related species, were studied for heterologous expression in E. coli. In this study, when an expressed tag sequence encoding signal peptide, His-S or GST-tag was fused to the 5′ end of LMW-GS or gliadin gene as the leading sequence, all recombination genes could be stably expressed at a high level. On the contrast, as expected, the inserted genes encoding mature protein failed without an expressed tag sequence. This result indicated that using expressed tag sequences as leading sequences could promote LMW-GS and gliadin genes to be well expressed in E. coli. Further transcriptional analysis by quantitative real-time PCR (qRT-PCR) showed transcription levels of recombination genes (e.g. GST-Glutenin, His-S-Glutenin and SP∗-His-Glutenin) were 4-fold to 33-fold higher than those of the LMW-GS genes, which suggested these expressed tag sequences might play an important role in stimulating transcription. The possible molecular mechanism under this phenomenon was discussed.
Details
- Title
- High-level expression of LMW-GS and α-gliadin genes promoted by the expressed tag sequence of 5′ end in Escherichia coli
- Authors/Creators
- P. Jiang (Author/Creator)K. Wang (Author/Creator)J. Gao (Author/Creator)X. Zheng (Author/Creator)J. Feng (Author/Creator)W. Ma (Author/Creator)Y. Yan (Author/Creator)X. Li (Author/Creator)
- Publication Details
- Protein Expression and Purification, Vol.105, pp.54-60
- Publisher
- Academic Press
- Identifiers
- 991005540582507891
- Copyright
- © 2014 Elsevier B.V.
- Murdoch Affiliation
- School of Veterinary and Life Sciences
- Language
- English
- Resource Type
- Journal article
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- 3 Agriculture, Environment & Ecology
- 3.85 Food Science & Technology
- 3.85.99 Cereal Starch Properties
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- Biochemical Research Methods
- Biochemistry & Molecular Biology
- Biotechnology & Applied Microbiology
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- Biology & Biochemistry