While polypeptides originating from ribosomes stalled in the pseudoknot varied only slightly in molecular weight, they varied significantly in their pI. Based on the 'Downstream Stop' construct, we calculated a theoretical 2D SDS-PAGE assay of a growing polypeptide as consecutive codons are translated (shown in Figure 2A ). At around 28 kDa, the trace splits into two, the triangles denote the non-frameshifted product and the circles denote the À1 frameshifted product. Red symbols denote codons inside the pseudoknot. Experimental data originating from the 'Downstream Stop' construct is shown in Figure 2B , the theoretically expected features are indeed present, e.g. both the non-frameshifted (DS-stop) and the À1 frameshifted (FS) products are visible. The heat shock proteins GroEL and DnaK serve as landmarks on the gel. Interestingly, a series of polypeptides originating from ribosomes stalled inside the pseudoknot appeared (inside dashed red line). For comparison, a standard SDS-PAGE of the same sample is shown in Figure 2C , here, the second level of information (isoelectric point) is lost and the relative blurry bands are difficult to interpret.