Abstract
Marine macroalgae play an important role in marine coastal ecosystems and are widely used as sea vegetation foodstuffs and for industrial purposes. Therefore, there have been increased demands for useful species and varieties of these macroalgae. However, genetic transformation in macroalgae has not yet been established. We have developed a dominant selection marker for stable nuclear transformation in the red macroalga Pyropia yezoensis. We engineered the coding region of the aminoglycoside phosphotransferase gene aph7″ from Streptomyces hygroscopicus to adapt codon usage of the nuclear genes of P. yezoensis. We designated this codon-optimized aph7″ gene as PyAph7. After bombarding P. yezoensis cells with plasmids containing PyAph7 under the control of their endogenous promoter, 1.9 thalli (or individuals) of hygromycin-resistant strains were isolated from a 10-mm square piece of the bombarded thallus. These transformants were stably maintained throughout the asexual life cycle. Stable expression of PyAph7was verified using Southern blot analysis and genomic PCR and RT-PCR analyses. PyAph7 proved to be a new versatile tool for stable nuclear transformation in P. yezoensis.Electronic supplementary materialThe online version of this article (doi:10.1007/s10126-013-9549-5) contains supplementary material, which is available to authorized users.
Highlights
The marine red macroalga Pyropia yezoensis is one of the most important marine crop
A laboratory culturing system in which the life cycle of P. yezoensis could be completed within a few months was established (Kuwano et al 1996)
As an initial step in establishing stable transformation, we previously developed a transient gene expression system to monitor gene expression in P. yezoensis cells using particle bombardment
Summary
The marine red macroalga Pyropia yezoensis (nori in Japanese) is one of the most important marine crop. In the present study, we synthesized a codonoptimized aph7′′ gene and examined its utility as a selection marker for stable nuclear transformation in P. yezoensis. Gametophytes of P. yezoensis strain TU-1 and transformants were cultured in enriched sea life (ESL) medium under conditions described by Fukuda et al (2008).
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