RNA complementary to the 5' UTR of mRNA triggers effective silencing in Saccharomyces cerevisiae.
Conditional silencing of target genes in Saccharomyces cerevisiae by antisense RNAs expressed in vivo has been challenged. The MFalpha1::lacZ fusion present in S. cerevisiae SF51-3 was chosen as a model target, and fragments of this gene were cloned in reverse orientation into the expression vector pYES2, bearing the GAL1 promoter. Among the different antisense constructs tested, only the one complementary to the 5' UTR of target mRNA featured effective silencing. Nevertheless, the expression in vivo of this antisense RNA could not be properly tuned by the absence or presence of galactose in the culture medium. Accordingly, conditional silencing could not be attained by this antisense hosted into pYES2. On the contrary, cloning the same antisense construct into the expression vector pSAL4 yielded a fully conditional silencing linked to the control of antisense expression by the absence or presence of Cu(2+) into the culture medium.
Bonoli
M
M
Department of Industrial Chemistry, University of Bologna, Viale Risorgimento 4, 40136 Bologna, Italy.
Graziola
M
M
Poggi
V
V
Hochkoeppler
A
A
D020121:5' Untranslated Regions; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D020868:Gene Silencing; D018390:Gene Targeting / Q000379:methods*; D015202:Protein Engineering / Q000379:methods; D016372:RNA, Antisense / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics*
D020121:5' Untranslated Regions; D016372:RNA, Antisense; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins