Regulatory elements in vectors containing the ctEF-1 alpha first intron and double enhancers for an efficient recombinant protein expression system

doi:10.1038/s41598-018-33500-0

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	Regulatory elements in vectors containing the ctEF-1 alpha first intron and double enhancers for an efficient recombinant protein expression system
	Lee, Chi-Pin 1; Ko, Albert Min-Shan 2; Chiang, Shang-Lun 1,3; Lu, Chi-Yu 4; Tsai, Eing-Mei 5; Ko, Yin-Chin 1
	2018-10-18
发表期刊	SCIENTIFIC REPORTS
ISSN	2045-2322
卷号	8 页码:9
通讯作者	Ko, Yin-Chin(ycko0406@gmail.com)
摘要	To establish a stable and scalable transient protein production system, we modified the EF-1 first intron size and verified the order of two recombinant enhancers downstream of the SV40 polyA sequence. This new vector was named pHH-Gemini (pHH-GM1) and was used to express alpha kinase 1 (ALPK1) and various other proteins, NLRP3, F-actin, Camodulin, PP2A, URAT1, Rab11a and myosin IIA. The results showed that, compared with six commercial plasmids, pHH-GM1 significantly enhanced His-HA-ALPK1 expression in a western blot analysis of transfected HEK293T cells. The expression of various other genes was also successful using the pHH-GM1 vector. In addition, we inserted turbo green florescence protein (tGFP) into the pHH-GM1 vector, and an improvement in fluorescence intensity was observed after transient transfection of HEK293T cells. For large-scale production, protein production was tested by standard supplementation with one volume of medium, and volumetric yields of 2 and 2.3 mg/L were achieved with pHH-GM1-ALPK1 in HEK293-F and CHO-S cells, respectively. We found that cell viability was more than 70%11 days after cells were transfected with the pHH-GM1 vector. The pHH-GM1 vector with the ctEF-la first intron and double enhancers, Simian virus 40 and Cytomegalovirus (SV40 and CMV) is an efficient CMV promoter-based gene expression system that can potentially be applied to study genes of interest and improve protein production.
DOI	10.1038/s41598-018-33500-0
关键词[WOS]	MAMMALIAN-CELLS ; INDUCTION ; GENE ; PROMOTER
收录类别	SCI
语种	英语
资助项目	Ministry of Science and Technology[MOST 105-2632-B-039-001] ; Ministry of Science and Technology[MOST 106-2314-B-039-017-MY3] ; China Medical University Hospital[DMR-107-108] ; China Medical University Hospital[DMR-107-208]
项目资助者	Ministry of Science and Technology ; China Medical University Hospital
WOS研究方向	Science & Technology - Other Topics
WOS类目	Multidisciplinary Sciences
WOS记录号	WOS:000447591000053
出版者	NATURE PUBLISHING GROUP
引用统计
文献类型	期刊论文
条目标识符	http://119.78.100.205/handle/311034/9015
专题	中国科学院古脊椎动物与古人类研究所
通讯作者	Ko, Yin-Chin
作者单位	1.China Med Univ, China Med Univ Hosp, Environm Omics Dis Res Ctr, Taichung 40402, Taiwan 2.Chinese Acad Sci, Key Lab Vertebrate Evolut & Human Origins, IVPP, Beijing 100044, Peoples R China 3.China Med Univ, Coll Publ Hlth, Dept Hlth Risk Management, Taichung 40402, Taiwan 4.Kaohsiung Med Univ, Coll Med, Dept Biochem, Kaohsiung 80708, Taiwan 5.Kaohsiung Med Univ, Coll Med, Grad Inst Med, Kaohsiung 80708, Taiwan
推荐引用方式 GB/T 7714	Lee, Chi-Pin,Ko, Albert Min-Shan,Chiang, Shang-Lun,et al. Regulatory elements in vectors containing the ctEF-1 alpha first intron and double enhancers for an efficient recombinant protein expression system[J]. SCIENTIFIC REPORTS,2018,8:9.
APA	Lee, Chi-Pin,Ko, Albert Min-Shan,Chiang, Shang-Lun,Lu, Chi-Yu,Tsai, Eing-Mei,&Ko, Yin-Chin.(2018).Regulatory elements in vectors containing the ctEF-1 alpha first intron and double enhancers for an efficient recombinant protein expression system.SCIENTIFIC REPORTS,8,9.
MLA	Lee, Chi-Pin,et al."Regulatory elements in vectors containing the ctEF-1 alpha first intron and double enhancers for an efficient recombinant protein expression system".SCIENTIFIC REPORTS 8(2018):9.