Content area
Full Text
PROTOCOL
High-efciency yeast transformation using the LiAc/SS carrier DNA/PEG method
R Daniel Gietz1 & Robert H Schiestl2
u
t
a
N 8
2 natureprotocols
/ m
o
1Department of Biochemistry and Medical Genetics, University of Manitoba, T250-770 Bannatyne Ave., Winnipeg, Manitoba R3E 0W3, Canada. 2Department of Pathology, Environmental Health and Radiation Oncology, UCLA School of Public Health and David Geffen School of Medicine, 650 Charles E. Young Drive South, Los Angeles, California 90095, USA. Correspondence should be addressed to R.H.S. ([email protected]).
Published online 31 January 2007; corrected online 20 November 2008 (details online); doi:10.1038/nprot.2007.13
Here we describe a high-efciency version of the lithium acetate/single-stranded carrier DNA/PEG method of transformation of Saccharomyces cerevisiae. This method currently gives the highest efciency and yield of transformants, although a faster protocol is available for small number of transformations. The procedure takes up to 1.5 h, depending on the length of heat shock, once the yeast culture has been grown. This method is useful for most transformation requirements.
INTRODUCTIONThe transformation of Saccharomyces cerevisiae with alkali cations was rst described in 1983 by Ito et al.1 Many improvements have taken place over the past 20 odd years, making the procedure more efcient in generating transformants as well as shortening the procedure. The most signicant improvement was the inclusion of single-stranded DNA as carrier, giving rise to an increase in transformation efciency2. This technique has been optimized for cell number, carrier DNA concentration and plasmid DNA concentration3. This transformation method can be modied and used for different purposes for yeast molecular biology4. The high-efciency transformation protocol can be employed to screen
various types of plasmid libraries. In addition, this protocol can be used to transform linear DNA constructs for DNA knockout experiments as well as oligonucleotides in yeast56. The various
S. cerevisiae transformation methods were recently reviewed7 and the reader should refer to that document for a history of yeast transformation methods. This protocol can also be used for transformation of other yeast species with some modication7.
If transforming a small number of transformants, a quicker and easier protocol is available8. Alternatively, the protocol can be scaled up using the library screen transformation protocol9 or a 96-well plate10.
c
.
e
r
u
t
a
n
.
w
w
w
/
/