Could anyone share their experience of transforming "cell wall" strains of Chlamydomonas? Literature suggests "autolysin" treatment to be the most preferred method followed by Biolistic gun for transforming cell wall strains.
However has anyone tried Electroporation or other methods?
Hope this reference would help. Physiol Plant. 2009 Jul;136(3):336-50.
Electroporation works pretty well on most strains; we regularly use this technique to transform chlamy
Hello Julie .. Thanks for your feedback.
Could you kindly share your experience with the same. What kind of instruments you use and what kind of strains you work with.
Hi Deepak,
We use a BTX Electroporation System Electro Cell Manipulator 600 with the following settings: 500V/Capacitance & resistance, R1 resistance timing, 200 + 800 in for capacitance timing (others out) and 200V voltage charge. We electroporate Chlamy in 1mm gap electroporation cuvettes. We routinely use this method for many strains with cell walls (e.g. wild-types 620 621 137c, IFT mutants, IFT rescues). It does help to linearize dna for transformation. Hope this helps!
Julie.. thank you for the feedback. I will try implementing the settings on the instrument we have and hope for the best..
can any help me to give setting for cell wall less Chlaymydomonas electroporation using bio rad gene pulser. Any inbuilt program will work?
Hi Rakesh,
In our experience in the lab, electroporation destroys a lot of cells and you end up recovering only few transformants. On the other hand, glass bead transformation works well and is predominantly used for transforming cell wall less strains.
Good luck
As Deepak said, electroporation is not needed for cell wall less strains.
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