Transgenesis in microalga Chlamydomonas reinhardtii: current approaches

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Abstract

Microalgae are a rich source of biologically active substances of natural origin, which have potential for use in pharmaceutical, agricultural, food and industrial production. Genetic engineering of microalgae opens up great prospects for creating improved strains that produce various food additives, commercial enzymes, as well as proteins for therapeutic purposes — antibodies, hormones and vaccines. Chlamydomonas reinhardtii P.A. Dang. is a unicellular green alga, a reference organism for studying the genetics of photosynthesis and developing new genetic engineering approaches in microalgae. The advantages of C. reinhardtii include the ability to transform all three of its genomes (nuclear, mitochondrial and chloroplast), low cost and ease of cultivation, safety for humans and the presence of a system for post-translational modification of proteins, which makes this organism a potential platform for use in biotechnology. Over the past few years, significant advances have been made in transgenesis of C. reinhardtii, including the use of new techniques based on the CRISPR/Cas9 genome editing technology. In this review, we summarize the available information on current approaches to transgenesis of the unicellular green alga C. reinhardtii: 1) general principles of transgenic constructs design for transformation of the nuclear and chloroplast genome, 2) popular selection markers used, 3) methods of cell transformation, 4) methods of genome editing using the CRISPR/Cas9 system.

About the authors

Pavel A. Virolainen

Saint Petersburg State University

Email: st085618@student.spbu.ru
ORCID iD: 0000-0001-5918-9395
SPIN-code: 6564-9350
Scopus Author ID: 57883811500

PhD student

Russian Federation, 7/9 Universitetskaya emb., Saint Petersburg, 199034

Elena M. Chekunova

Saint Petersburg State University

Author for correspondence.
Email: e.chekunova@spbu.ru
ORCID iD: 0000-0001-8942-4771
SPIN-code: 2788-6386
Scopus Author ID: 6701797455

Dr. Sci. (Biology), Senior Teacher

Russian Federation, 7/9 Universitetskaya emb., Saint Petersburg, 199034

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2. Fig. 1. Overall structure of transgenic constructs for transformation of the nuclear and chloroplast genome of C. reinhardtii: a — construct for transformation of the nuclear genome; b — construct for transformation of the chloroplast genome. ПГ — homology arms; П — promoter/5'-untranslated region; И — intron; T — terminator/3'-untranslated region; ПП — direct repeats. Additional explanations are given in the text

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3. Fig. 2. Strategies for delivering components of the CRISPR/Cas9 system into C. reinhardtii cells: a — integrative system; b — episomal system; c — ribonucleoprotein system. Additional explanations are given in the text

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