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Efficient selection of phleomycin-resistant Saccharomyces cerevisiae transformants
T J Wenzel1, A Migliazza, H Y Steensma
1Department of Cell Biology and Genetics, Leiden University, The Netherlands.
Yeast (Chichester, England)
|August 1, 1992
Summary
The yeast marker Tn5ble provides phleomycin resistance in Saccharomyces cerevisiae. Optimal transformation frequencies are achieved with a 6-hour incubation in non-selective medium before selection.
Area of Science:
- Molecular Biology
- Yeast Genetics
- Biotechnology
Background:
- The Tn5ble marker confers phleomycin resistance in yeast.
- Transformation efficiency is influenced by pre-selection incubation periods.
Purpose of the Study:
- To optimize transformation frequencies using the Tn5ble marker in Saccharomyces cerevisiae.
- To determine the critical incubation period for maximizing phleomycin-resistant transformants.
Main Methods:
- Transformation of Saccharomyces cerevisiae with the Tn5ble marker.
- Incubation in non-selective medium for varying durations.
- Selection for phleomycin resistance.
- Comparison with uracil prototrophy selection.
Main Results:
- A 6-hour incubation period yielded optimal transformation frequencies.
- Frequencies reached up to 10(5) transformants per microgram of plasmid.
- These frequencies were comparable to those obtained with uracil prototrophy selection.
Conclusions:
- The Tn5ble marker is an effective dominant yeast marker for phleomycin resistance.
- A specific pre-selection incubation period is critical for maximizing transformation efficiency.
- The optimized protocol offers a viable alternative for yeast genetic manipulation.