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Expression levels of domestic cDNA cassettes integrated in the nuclear genomes of various Chlamydomonas reinhardtii strains

Paper ID Volume ID Publish Year Pages File Format Full-Text
20476 43176 2014 4 PDF Available
Title
Expression levels of domestic cDNA cassettes integrated in the nuclear genomes of various Chlamydomonas reinhardtii strains
Abstract

We attempted to overexpress three types of expression cassettes, each of which contained a different open reading frame (ORF) of domestic Chlamydomonas cDNAs. Each ORF was strongly driven by an artificial hybrid promoter. We used two wild-type Chlamydomonas strains (i.e., CC-124 and CC-125) and two mutant strains [i.e., UV-mutated (UVM) 4 and UVM11] that have been reported to have a high potency for expressing nondomestic nuclear transgenes. We found that the 1-deoxy-d-xylulose-5-phosphatesynthase (DXS1), 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR1), and squalene synthase (SQS) cassettes were not readily overexpressed in the wild-type strains at levels where the products were clearly detectable by Western blotting using a monoclonal antibody. In contrast, Western blot-positive SQS cassette transformants were frequently detected in the UVM4 and UVM11 strains, i.e., at an approximately 4.5 times higher frequency than that in the CC-124 wild-type strain. Moreover, transformants that accumulated large amounts of the SQS protein were obtained frequently in the UVM4 and UVM11 strains, i.e., the frequency was approximately 2.2 times higher than that in the CC-124 strain. However, a position effect of the integrated expression cassette was obviously detected not only in the wild-type but also in UVM strains. This suggests that the epigenetic repression mechanism of transgenic genes was not completely knocked out, even in the UVM strains. Further improved Chlamydomonas strains are essential to facilitate high-throughput screening of transformants that express nuclear transgenes at a high level.

Keywords
cDNA expression cassettes; Chlamydomonas reinhardtii; Nuclear transformation; UV-mutated strains; Overexpression
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Expression levels of domestic cDNA cassettes integrated in the nuclear genomes of various Chlamydomonas reinhardtii strains
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 117, Issue 5, May 2014, Pages 613–616
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us