Engineering of an endogenous phytoene desaturase gene as a dominant selectable marker for Chlamydomonas reinhardtii transformation and enhanced biosynthesis of carotenoids

被引：69
|
作者：
Liu, Jin [1 ,4 ]
Gerken, Henri [2 ]
Huang, Junchao [3 ]
Chen, Feng [4 ]
机构：
[1] Univ Maryland, Ctr Environm Sci, Inst Marine & Environm Technol, Baltimore, MD 21201 USA
[2] Arizona State Univ, Dept Appl Sci & Math, Mesa, AZ USA
[3] Chinese Acad Sci, Kunming Inst Bot, Kunming, Peoples R China
[4] Peking Univ, Coll Engn, Inst Food & Bioresource Engn, Beijing 100871, Peoples R China
来源：
PROCESS BIOCHEMISTRY | 2013年 / 48卷 / 5-6期
关键词：
Chlamydomonas reinhardtii; Carotenoids; Norflurazon; Phytoene desaturase; Selectable marker; Site-directed mutation; NUCLEAR TRANSFORMATION; FOREIGN GENE; EXPRESSION; RESISTANCE; NORFLURAZON; MUTANTS; PCR;
D O I：
10.1016/j.procbio.2013.04.020
中图分类号：
Q5 [生物化学]; Q7 [分子生物学];
学科分类号：
071010 ; 081704 ;
摘要：
A phytoene desaturase (PDS) gene was cloned and characterized from the unicellular green microalga Chlamydomonas reinhardtii. Functional complementation analysis revealed C reinhardtii PDS (CrPDS) catalyzes the conversion of phytoene to the colored carotenoid zeta-carotene. A single amino acid substitution, L505F, enhanced its desaturation activity by 29%, as indicated by an in vitro enzymatic assay. In addition, CrPDS-L505F exhibited 27.7-fold higher resistance to the herbicide norflurazon. Glass bead-mediated delivery displayed a high transformation efficiency of C reinhardtii with CrPDS-L505F, demonstrating clearly that the engineered endogenous CrPDS is a dominant selectable marker for C. reinhardtii and possibly for other green algae. Furthermore, the expression of PDS could enhance the intracellular carotenoid accumulation of transformants, opening up the possibility of engineering the carotenogenic pathway for improved carotenoid production in microalgae. (C) 2013 Elsevier Ltd. All rights reserved.
引用
收藏
页码：788 / 795
页数：8
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