A transgene expression system for the marine microalgae Aurantiochytrium sp. KRS101 using a mutant allele of the gene encoding ribosomal protein L44 as a selectable transformation marker for cycloheximide resistance

• Published in: Bioprocess and biosystems engineering Vol. 36; no. 9; pp. 1191 - 1197
• Main Authors: Hong, Won-Kyung, Heo, Sun-Yeon, Oh, Baek-Rock, Kim, Chul Ho, Sohn, Jung-Hoon, Yang, Ji-Won, Kondo, Akihiko, Seo, Jeong-Woo
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer Berlin Heidelberg 01.09.2013; Springer Nature B.V
• Subjects: Algae; Alleles; Antifungal Agents - pharmacology; Biosynthesis; Biotechnology; Chemistry; Chemistry and Materials Science; Cycloheximide - pharmacology; Drug Resistance - drug effects; Drug Resistance - genetics; Environmental Engineering/Biotechnology; Fatty acids; Food Science; Gene Expression; Genetic Markers; Industrial and Production Engineering; Industrial Chemistry/Chemical Engineering; Microalgae; Mortierella alpina; Mutation; Original Paper; Proteins; Ribosomal Proteins - biosynthesis; Ribosomal Proteins - genetics; Stramenopiles - genetics; Stramenopiles - metabolism; Transgenes; Transgenic plants; Ribosomal protein; Cycloheximide resistance; Gene transformation
• ISSN: 1615-7591; 1615-7605
• DOI: 10.1007/s00449-012-0846-6

In the present study, we established a genetic system for manipulating the oleaginous heterotrophic microalgae Aurantiochytrium sp. KRS101, using cycloheximide resistance as the selectable marker. The gene encoding ribosomal protein L44 (RPL44) of Aurantiochytrium sp. KRS101 was first identified and characterized. Proline 56 was replaced with glutamine, affording cycloheximide resistance to strains encoding the mutant protein. This resistance served as a novel selection marker. The gene encoding the Δ12-fatty acid desaturase of Mortierella alpina , used as a reporter, was successfully introduced into chromosomal DNA of Aurantiochytrium sp. KRS101 via 18S rDNA-targeted homologous recombination. Enzymatic conversion of oleic acid (C18:1) to linoleic acid (C18:2) was detected in transformants but not in the wild-type strain.