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Gene silencing by artificial microRNAs in Chlamydomonas
Author(s) -
Zhao Tao,
Wang Wei,
Bai Xue,
Qi Yijun
Publication year - 2009
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/j.1365-313x.2008.03758.x
Subject(s) - chlamydomonas reinhardtii , chlamydomonas , biology , gene silencing , gene , genetics , microrna , genome , computational biology , mutant , microbiology and biotechnology
Summary Chlamydomonas reinhardtii is a unicellular green alga. It is a model system for studying functions of the chloroplast, basal body and flagella. The completion of the Chlamydomonas genome sequence makes it possible to use reverse genetic approaches in this organism. Chlamydomonas contains a set of endogenous microRNAs (miRNAs) that down‐regulate their target gene expression through mRNA cleavage. Here we developed an artificial miRNA‐based strategy to knock down gene expression in Chlamydomonas . Using an endogenous Chlamydomonas miRNA precursor as the backbone, we constructed two artificial miRNAs (amiRNAs) targeting the MAA7 and RBCS1 / 2 genes, respectively. When overexpressed, these two amiRNAs could cleave their respective targets precisely at the predicted sites, resulting in greatly decreased accumulation of MAA7 and RBCS1 / 2 transcripts and expected mutant phenotypes. We further showed that the two amiRNAs could be produced simultaneously from a dimeric amiRNA precursor. We anticipate that the amiRNA technology developed in this study will be very useful in assessing the functions of individual genes and in genome‐wide approaches.