Codon deoptimization away from frq leads to untimely transcription termination.
(A) Northern blot showing the presence of truncated frq mRNA species in both de-optimized strains using an RNA probe targeted to 5' end of frq mRNA (indicated in ; Figure 2-figure supplement 1E). * indicates a non-specific band. (B) Northern blot showing both full-length and truncated frq mRNA are enriched in poly(A)-containing RNAs. Equal amounts of total RNA or poly(A) RNA were loaded in each lane. Probe specific for 5' end of frq was used. (C) Poly(A) sites mapped by 3' RACE. Arrows indicate the mapped poly(A) sites, the red arrows indicate the major poly(A) site that was found in both frq-deopt1 and frq-deopt2 strains, and the black line indicates potential PAS motif (AUAAAU in frq-deopt1 and AAUAAA in frq-deopt2). Nucleotides that are mutated are shown in red. (D) ChIP assay showing RNA http://datingranking.net/introvert-dating/ pol II levels at the frq transgene loci in the wt-frq-aq and frq-deopt2-aq strains. The ChIP results were normalized by input DNA and represented as Input%.