FIG. 2.

Selective identification of expanded TNR. (A) Sequences represent the TNR sequences used for this study. Nomenclature is that of the lagging daughter strand. (B) Genetic assay to select for TNR expansions in yeast. The region controlling expression of the reporter gene URA3 is shown, including the TATA box, the TNR region, an out-of-frame initiation codon, the preferred transcription initiation site (I), and the start of the URA3 gene. The upper diagram demonstrates the starting construct. The brackets represent a window of potential transcription initiation sites located 55 to 125 bp from the TATA box. With a 25-TNR tract, the transcription window includes the preferred site I, thus leading to expression of URA3 and making the yeast 5FOA sensitive. The lower diagram demonstrates what happens when the TNR tract expands to ≥30 repeats. The bracketed window does not include site I. Transcription initiating 5′ of the preferred initiation site will include the out-of-frame ATG, resulting in translational incompetence and therefore resistance to 5FOA.