Figure 6

Results of linker library screening. (A) Histogram of FRET ratio changes of all colonies screened from lib2. The average FRET change in colonies was 14% with a standard deviation of 4%. (B) Linker combinations identified during screening of lib2. The horizontal and the vertical axes represent the number of amino acids constituting the first linker (L1) and the third linker (L3), respectively. The inputs for the second linker (L2) are sequences of 0, 2, 4, 6, 8, 10, 14, and 20 residues in length. Identified L1 and L3 combinations are indicated on the grid by coloured circles the size of which represents the number of sequenced clones with a given combination of L1 and L3, while the number of amino acids constituting L2 of these clones is indicated by the numerical values assigned to the circles. Linker combinations exhibiting the highest FRET ratio changes (> 20%) are represented by the green circle; orange circles represent clones with average-to-low ratio changes (< 20%); and grey circles represent clones picked at random. (C) Data histogram of FRET ratio changes of colonies screened from lib3. The average FRET change in colonies was 38% with a standard deviation of 16%. (D) Linker combinations identified during screening of lib3. Input for L2 are linkers of 6, 8, 10, 14, 15, 20, 25, or 30 residues in length. Linker combinations exhibiting high FRET ratio changes (> 70%) are represented with green circles and those showing average-to-low ratio changes (< 70%) are represented by orange circles.