The extent of silencing varied in the different transformants between high and moderate levels.
In 24 h of transfection, transfected cells were sorted to purify the different transformants by fluorescence activated cell sorter (FACS).
Glycerol stocks of spores of the different transformants made on SFM medium were precisely titrated by plating different spore dilutions on HT agar containing 50 μg.ml−1 thiostrepton.
The stability of the resistance phenotype varied considerably among the different transformants (data not shown).
The different transformants also exhibited distinct growth rates in liquid media lacking drugs.
The amount of As III) taken up by the different transformants corresponded well with the degree of As III) sensitivity seen when growing the respective transformant on As III -containing medium (rAQP9 > Atnip5 1Δ2 67 > Atnip6 1Δ2 69 > Atnip7 1Δ2 35 ≈ AtNIP7 1; see Figure 2A and 2B).
The different transformant strains were produced in different labs at different times, and have different wild-type backgrounds.
A wide variation in heterologous gene expression in S. cerevisiae is not unusual when episomal 2 μ ori-based plasmids are used, possibly because of variation in the plasmid copy number between different transformants [ 33, 34].
Thus, even the extreme variability in luciferase activity between different transformants was permanent.
Figure 2 Correlation between the level of transcription of the aphII gene and the survival rates of different transformants in the presence of 100 μg.ml -1 neomycin.
