TY  - JOUR
TI  - Multiple Modification of Chromosomal Loci Using URA5.3 Selection Marker in the Unicellular Red Alga Cyanidioschyzon merolae
AU  - Takemura, Tokiaki  
AU  - Imamura, Sousuke  
AU  - Kobayashi, Yuki  
AU  - Tanaka, Kan  
VL  - 9
IS  - 7
PY  - 2019
DA  - 2019/04/05
SP  - e3204
C1  - Bio-protocol 2019;9:e3204
DO  - 10.21769/BioProtoc.3204
UR  - https://doi.org/10.21769/BioProtoc.3204
AB  - The unicellular red alga Cyanidioschyzon merolae has been used as a eukaryotic photosynthetic model for various basic and applied studies. Although the nuclear genome of C. merolae can be modified by homologous recombination with exogenously introduced DNA, it has been difficult to modify multiple chromosome loci within the same strain because of the limited number of available positive selection markers. Recently, we reported a modified URA5.3 gene cassette (URA5.3T), which can be used repeatedly for nuclear genome transformation using the pMKT plasmid vectors for epitope tagging (3x FLAG- or 3x Myc-) of nuclear-encoded proteins. In addition, these plasmid vectors can also be used to knock out multiple genes one by one. This report describes the construction of DNA fragments for transformation and the detailed transformation procedure.
KW  - Cyanidioschyzon merolae
KW  - Epitope tagging
KW  - Gene knocking out
KW  - Homologous recombination
KW  - Selectable marker recycling
KW  - URA5.3
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False