Protoplast Transformation: Difference between revisions
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*[[P buffer]] | *[[P buffer]] | ||
*1 g polyethylene glycol 1000 (PEG) | *1 g polyethylene glycol 1000 (PEG) | ||
*[[R5]] or [[R2YE]] plate | *[[R5]] or [[R2YE]] plate | ||
| Line 12: | Line 11: | ||
#Add 200 µl PEG-P buffer mixture and a DNA solution to 50 µl of the protoplast suspension. | #Add 200 µl PEG-P buffer mixture and a DNA solution to 50 µl of the protoplast suspension. | ||
#Plate it onto [[R5]] or [[R2YE]]. | #Plate it onto [[R5]] or [[R2YE]]. | ||
#Incubate the plate at | #Incubate the plate at 28-30ºC overnight (typically 12-18 hours). | ||
#Overlay 1 ml sterilised water containing nalidixic acid and an appropriate antibiotic. | #Overlay 1 ml sterilised water containing nalidixic acid and an appropriate antibiotic. | ||
Latest revision as of 05:26, 31 March 2020
Materials
Instructions
- Follow the instruction in Protoplasts Formation, Regeneration and Transformation
- Add 3.85 ml P buffer to 1 g PEG sterilised.
- Add 200 µl PEG-P buffer mixture and a DNA solution to 50 µl of the protoplast suspension.
- Plate it onto R5 or R2YE.
- Incubate the plate at 28-30ºC overnight (typically 12-18 hours).
- Overlay 1 ml sterilised water containing nalidixic acid and an appropriate antibiotic.
