Tools for chloroplast transformation in Chlamydomonas: expression vectors and a new dominant selectable marker.
Reverse-genetic studies of chloroplast genes in the green alga Chlamydomonas reinhardtii have been hampered by the paucity of suitable selectable markers for chloroplast transformation. We have constructed a series of vectors for the targeted insertion and expression of foreign genes in the Chlamydomonas chloroplast genome. Using these vectors we have developed a novel selectable marker based on the bacterial gene aphA-6, which encodes an aminoglycoside phosphotransferase. The aphA-6 marker allows direct selection for transformants on medium containing either kanamycin or amikacin. The marker can be used to inactivate or modify specific chloroplast genes, and can be used as a reporter of gene expression. The availability of this marker now makes possible the serial transformation of the chloroplast genome of Chlamydomonas.
Bateman
J M
JM
Department of Biology, University College London, UK.
Purton
S
S
D000583:Amikacin / Q000378:metabolism; D000818:Animals; D015139:Blotting, Southern; D002696:Chlamydomonas / Q000235:genetics*; D002736:Chloroplasts / Q000235:genetics*; D004352:Drug Resistance, Microbial / Q000235:genetics; D004578:Electron Spin Resonance Spectroscopy; D018390:Gene Targeting; D005819:Genetic Markers; D019868:Kanamycin Kinase / Q000235:genetics; D007613:Kanamycin Resistance / Q000235:genetics; D008957:Models, Genetic; D016254:Mutagenesis, Insertional; D010957:Plasmids; D014170:Transformation, Genetic
D005819:Genetic Markers; D000583:Amikacin; D019868:Kanamycin Kinase