Hi, everybody!
Now I am trying to transform wild type C.reinhardtii CC124 with 1302 vector by glass bead. I am not so familiar with Chlamydomonas strains. In research papers, it was reported that mating type (+) and (-) was mixed before transformation and they obtained mating type (+) and (-) from from stock center . I have only wild type C.reinahrdtii CC124. To transform by glass bead, how should I continue with this wild type strain? I am strongly expecting everybody's suggestion and will treasure any advice.
The glass bead method is only suitable for transformation of cell wall mutants of Chlamydomonas. See the link below for the original paper. As your strain appears to not have the cell wall mutation you will probably need to use a biolistic transformation protocol. The mating types are only necessary if you are aiming to then mate your cells to get multiple different transgenes into your cells. The vegetative cells are monoploid so it is not necessary to generate crosses to get homozygotes. It is probably easier to transform your transformed cells with your second vector to generate double transgenes.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC53444/?page=1
Dear Ellis C O'Neill,
Thank you so much for kind suggestion and sharing your knowledge. I have got invaluable knowledge.I fully hope that your suggestion will support my research work.
You only need to transform using particle bombardment if you're trying to get DNA into the chloroplast. You can also use electroporation for nuclear transformations. Also, mating will not help you introduce multiple mutations into the chloroplast because it is only maternally inherited.
Dear James Gregory,
Thank you so much for your answer and suggestion. For me, I have got invaluable knowledge to continue my research.
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