DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins

Kwangryul Baek; Duk Hyoung Kim; Jooyeon Jeong; Sang Jun Sim; Anastasios Melis; Jin Soo Kim; Eonseon Jin; Sangsu Bae

doi:10.1038/srep30620

DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins

Kwangryul Baek, Duk Hyoung Kim, Jooyeon Jeong, Sang Jun Sim, Anastasios Melis, Jin Soo Kim, Eonseon Jin, Sangsu Bae

Department of Chemical and Biological Engineering

Research output: Contribution to journal › Article › peer-review

202 Citations (Scopus)

Abstract

Microalgae are versatile organisms capable of converting CO 2, H 2 O, and sunlight into fuel and chemicals for domestic and industrial consumption. Thus, genetic modifications of microalgae for enhancing photosynthetic productivity, and biomass and bio-products generation are crucial for both academic and industrial applications. However, targeted mutagenesis in microalgae with CRISPR-Cas9 is limited. Here we report, a one-step transformation of Chlamydomonas reinhardtii by the DNA-free CRISPR-Cas9 method rather than plasmids that encode Cas9 and guide RNAs. Outcome was the sequential CpFTSY and ZEP two-gene knockout and the generation of a strain constitutively producing zeaxanthin and showing improved photosynthetic productivity.

Original language	English
Article number	30620
Journal	Scientific reports
Volume	6
DOIs	https://doi.org/10.1038/srep30620
Publication status	Published - 2016 Jul 28

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title = "DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins",

abstract = "Microalgae are versatile organisms capable of converting CO 2, H 2 O, and sunlight into fuel and chemicals for domestic and industrial consumption. Thus, genetic modifications of microalgae for enhancing photosynthetic productivity, and biomass and bio-products generation are crucial for both academic and industrial applications. However, targeted mutagenesis in microalgae with CRISPR-Cas9 is limited. Here we report, a one-step transformation of Chlamydomonas reinhardtii by the DNA-free CRISPR-Cas9 method rather than plasmids that encode Cas9 and guide RNAs. Outcome was the sequential CpFTSY and ZEP two-gene knockout and the generation of a strain constitutively producing zeaxanthin and showing improved photosynthetic productivity.",

author = "Kwangryul Baek and Kim, {Duk Hyoung} and Jooyeon Jeong and Sim, {Sang Jun} and Anastasios Melis and Kim, {Jin Soo} and Eonseon Jin and Sangsu Bae",

note = "Funding Information: This work was supported by grants from Institute for Basic Science (IBS-R021-D1) to J.-S.K.The Korea CCS R&D Center (KCRC) (NRF-2014M1A8A1049273) to E.J. and the Plant Molecular Breeding Center of Next Generation BioGreen 21 Program (PJ01119201) to S.B.",

year = "2016",

month = jul,

day = "28",

doi = "10.1038/srep30620",

language = "English",

volume = "6",

journal = "Scientific Reports",

issn = "2045-2322",

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T1 - DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins

AU - Baek, Kwangryul

AU - Kim, Duk Hyoung

AU - Jeong, Jooyeon

AU - Sim, Sang Jun

AU - Melis, Anastasios

AU - Kim, Jin Soo

AU - Jin, Eonseon

AU - Bae, Sangsu

N1 - Funding Information: This work was supported by grants from Institute for Basic Science (IBS-R021-D1) to J.-S.K.The Korea CCS R&D Center (KCRC) (NRF-2014M1A8A1049273) to E.J. and the Plant Molecular Breeding Center of Next Generation BioGreen 21 Program (PJ01119201) to S.B.

PY - 2016/7/28

Y1 - 2016/7/28

N2 - Microalgae are versatile organisms capable of converting CO 2, H 2 O, and sunlight into fuel and chemicals for domestic and industrial consumption. Thus, genetic modifications of microalgae for enhancing photosynthetic productivity, and biomass and bio-products generation are crucial for both academic and industrial applications. However, targeted mutagenesis in microalgae with CRISPR-Cas9 is limited. Here we report, a one-step transformation of Chlamydomonas reinhardtii by the DNA-free CRISPR-Cas9 method rather than plasmids that encode Cas9 and guide RNAs. Outcome was the sequential CpFTSY and ZEP two-gene knockout and the generation of a strain constitutively producing zeaxanthin and showing improved photosynthetic productivity.

AB - Microalgae are versatile organisms capable of converting CO 2, H 2 O, and sunlight into fuel and chemicals for domestic and industrial consumption. Thus, genetic modifications of microalgae for enhancing photosynthetic productivity, and biomass and bio-products generation are crucial for both academic and industrial applications. However, targeted mutagenesis in microalgae with CRISPR-Cas9 is limited. Here we report, a one-step transformation of Chlamydomonas reinhardtii by the DNA-free CRISPR-Cas9 method rather than plasmids that encode Cas9 and guide RNAs. Outcome was the sequential CpFTSY and ZEP two-gene knockout and the generation of a strain constitutively producing zeaxanthin and showing improved photosynthetic productivity.

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DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins

Abstract

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