Transformation with ADE2

Guillaume Stahl stahl at infobiogen.fr
Tue Jun 3 18:07:50 EST 1997


> ade2-101 strains?

ade-101 is an ochre mutation !

How is you negative control ? I used to transform trp1-289 (amber), and 
the background of reversion is quite high.
Furthermore, there are also potential extragenic mutation which would 
supress this allele (tRNAs, and of course omnipotent supressors like 
SUP35=eRF3; SUP45=eRF3,...).

> When I pick these small colonies and streak them out onto fresh plates,

Small colonies after transformation are often rho- clones (but then this 
phenotype should not reverse...). Test this on glycerol plates (Anyway, 
it'a a good idea to streak your yeast strains from time to time on this 
media, to get an homogenous rho+ strain)

Perhaps the explanation of the instability of your [Ade+] pnenotype is 
of prionic order : SUP35 may be a prion for yeast, and the prionic fomr, 
known as psi is an omnipotent suprressor, but without mendelian 
segregation...

Don't know if this helps or a contrario ;-)
Guillaume Stahl, France



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