Table 1.
Characteristics of the reference sequences used in this work.
Figure 1.
Percentage of probes that occur more than once in the raw library L.
Fraction of repeated probes in the library (blue bullets, left-hand scale) and total number of probes |Ll| (red boxes, right-hand scale) as a function of probe length l in a) HIV reverse transcriptase and b) HCV E1. L′ is the reduced set of unique sequences as explained in Methods (See Equation 3). Lines are a guide to the eye, and it should be noted that the right hand scale is logarithmic.
Figure 2.
Relationship between variability and the number of targets found.
Average fraction of targets 〈tf〉 found as a function of a) the number of reference sequences NRef used and b) average relative variability of the library calculated for probe length l of 15 nt. Selected target sequences are for genes in HCV: E1 (bullets), core (boxes), and ns4A (triangles) and Influenza np5 (circles), NCBI∶AY746942, NCBI∶AY365214, NCBI∶D84264 and NCBI∶CY000172, respectively. The error bars in part a) represent the standard error in the mean and are the same for b). Lines connecting the symbols are guides to the eye.
Figure 3.
Effect of filtering on targets found.
a) Number of fragments 〈tf〉 found the selected target sequence Influenza np5 (NCBI∶CY000076) that are correctly identified as a function of probe length l. The average standard error is 0.23% and was consistent across all the data points. b) The average number of probes 〈|Ll|〉 in the library as a function of probe length for each of the filtered libraries. The diamonds represent the library before applying the filtering. Remaining points represent the libraries where all sequences with GC content above 80% have been removed and thermodynamic filtering has been applied as described in Methods, with Δω = 1 (bullets), 0.5 (boxes), 0.2 (triangles) and 0 (circles). The average standard error was 9.5% of the library size and was consistent across all the data points. Lines are a guide to the eye.
Figure 4.
Equivalence parameters of probes in libraries and targets.
a) Equivalence parameter ω of probes in the library (L12 and NRef = 5) as a function of the sequence position m of the probe in the target sequence (np5). b) The equivalence parameter ω of probes that intersect the library and an unseen target T12(NCBI∶CY 000172) as a function of the sequence position m of the probe in the target sequence. The equivalence parameter ω is defined in eqn. 11 The beginning (m) and end (m+12) of the probe is shown by short up and down lines, respectively. The colour coding is: black for Δω>1, red for 0.5<Δω≤1, blue for 0.2<Δω≤0.5, green for 0<Δω≤0.2 and magenta for the most frequent equivalence parameter ωf or Δω = 0.
Figure 5.
Percentage coverage after filtering.
Average coverage by probes of gene positions 〈C〉 as a function of probe length l for Influenza H3N2 np5. The diamonds represent the unfiltered library and the remaining symbols represent the libraries filtered as described in Methods. Values for Δω are 1 (bullets), 0.5 (boxes), 0.2 (triangles) and 0 (circles). Lines are a guide to the eye. The average standard error is 0.5% and was consistent across all the data points.
Table 2.
Optimal Probe lengths for genes studied.