A method for the re-isolation of an autonomously replicating plasmid from Aspergillus transformants.
An autonomously replicating plasmid is expected to increase the frequency of Aspergillus transformation. To construct this type of plasmid, we developed a rapid method of re-isolating autonomously replicating plasmids from Aspergillus transformants. Transformants grown in MM medium under selective pressure for 1-2 days were converted to protoplasts with a cell wall lytic enzyme (e.g. Yatalase). The protoplasts were lysed with phenol/chloroform followed by precipitation with ethanol. The total DNA was treated with RNaseA, re-precipitated with PEG, and then used to transform E. coli. These re-isolated plasmids were mainly the plasmid monomer.
Ozeki
K
K
General Research Laboratory, Ozeki Corporation, Hyogo, Japan.
Hizume
K
K
Kanda
A
A
Hamachi
M
M
Nunokawa
Y
Y
D001230:Aspergillus / Q000235:genetics*; D004261:DNA Replication; D004274:DNA, Recombinant / Q000096:biosynthesis; D010957:Plasmids
D004274:DNA, Recombinant