Comparison between polyethylene glycol- and polyethylenimine-mediated transformation of Aspergillus nidulans.
Genetic transformation of many filamentous fungi is carried out by a protocol that utilizes polyethylene glycol (PEG) and calcium ion (Ca2+). This method has remained practically unchanged for more than 20 years, but the roles these molecules play are not definitively understood. To gain a better understanding, we have compared PEG transformation to a protocol using polyethylenimine (PEI) that is the basis for non-viral transfection in mammals and which has a well established molecular model for assisting DNA uptake. Protoplasts of Aspergillus nidulans could be transformed in the presence of Ca2+ with a relatively high ratio of PEI to DNA molecules. By comparing PEI and PEG in terms of interaction with DNA, fungal protoplasts, and response to different transformation conditions, we propose that the role of PEG is most likely to function after transforming DNA is incorporated into protoplasts, rather than the accepted view that it functions outside of the cell. Confirmation that protoplast fusion was not involved in DNA uptake is consistent with this hypothesis.
Kuwano
Tetsuya
T
Department of Biological Sciences, Faculty of Science, Shinshu University, Matsumoto, Nagano 390-8621, Japan.
Shirataki
Chikako
C
Itoh
Yasuo
Y
D001233:Aspergillus nidulans / Q000235:genetics*; D002118:Calcium / Q000378:metabolism; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000302:isolation & purification; D011092:Polyethylene Glycols / Q000737:chemistry*; D011094:Polyethyleneimine / Q000737:chemistry*; D011108:Polymers / Q000737:chemistry*; D011523:Protoplasts; D014170:Transformation, Genetic
D004271:DNA, Fungal; D011108:Polymers; D011092:Polyethylene Glycols; D011094:Polyethyleneimine; D002118:Calcium