NDLI: Cloning of a novel endogenous promoter for foreign gene expression in Phaeodactylum tricornutum

Content Provider	Springer Nature Link
Author	Lee, Eun Ha Song, Dae Geun Huda, Md Nazmul Shin, Bok Kyu Kim, Sang Min Kim, Da Hye Pan, Cheol Ho Erdene Ochir, Erdenedolgor Kim, Yoon Mi
Copyright Year	2016
Abstract	Phaeodactylum tricornutum is a model diatom, and its genomic sequence data and expressed sequence tag databases are available. This study was to discover a new endogenous promoter that drives strong constitutive expression of a protein of interest in P. tricornutum. To find promoter candidates, the intracellular proteins of P. tricornutum grown to stationary phase were extracted and identified by LC–MS/MS. Glutamine synthetase (GLNA) was one of the most abundantly expressed proteins during the stationary phase. Promoter is usually located on 5′ upstream region of open reading frame of the gene. Thus, two fragments of 5′ upstream region of the GLNA gene as putative promoters, 501 and 997 bp long, were amplified and cloned into enhanced Green Fluorescent Protein (eGFP) reporter systems. The constructed reporter systems were transformed into P. tricornutum and the eGFP expression levels were compared to those of reporter systems using the promoters of fcpA (fucoxanthin chlorophyll a/c binding protein A) and CIP1 (putative replication-associated proteins of a Chaetoceros lorenzianus-infecting DNA virus) as controls. The expression of eGFP driven by either GLNA promoter (501 and 997 bp) was linearly related to cell density, and eGFP was expressed constitutively regardless of the cultivation phase. The eGFP expression level driven by the GLNA promoters was at least 4 times higher than the fcpA-driven eGFP expression level at the stationary phase. The 501 and 997 bp regions of the GLNA promoter had similar activity patterns for transcribing the downstream gene. These results indicate that at least 501-bp region of the GLNA promoter can be used as a strong constitutive promoter in genetic engineering of P. tricornutum.
Starting Page	861
Ending Page	867
Page Count	7
File Format	PDF
ISSN	24680834
Journal	Journal of the Korean Society for Applied Biological Chemistry
Volume Number	59
Issue Number	6
e-ISSN	24680842
Language	English
Publisher	Springer Netherlands
Publisher Date	2016-11-18
Publisher Place	Dordrecht
Access Restriction	Subscribed
Subject Keyword	Glutamine synthetase promoter Transformation Phaeodactylum tricornutum Applied Microbiology Biological Techniques Constitutive expression Bioorganic Chemistry
Content Type	Text
Resource Type	Article
Subject	Organic Chemistry Biochemistry, Genetics and Molecular Biology

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