Description: SynaHybridise™ is an ungapped, non-heuristic oligonucleotide sequence verification tool, designed to validate microarray probe specificity and uniqueness.
Application Example: Ungapped, non-heuristical weight matrix search offers improved sensitivity in finding targets with multiple mismatches within a short oligonucleotide sequence. Linear weighted search penalizes all mismatches equally, while centre weighted uses lower penalties for mismatches near the probe ends than for mismatches near the centre. Centre weighted searches are designed to be consistent with reports and observations that indicate sequences with mismatches near the end of a probe may still hybridise and produce a false positive signal. Common heuristical search tools using fixed k-mer seeds to search for sequences that potentially bind to microarray probes have low sensitivity and specificity. Therefore by using SynaHybridise, centre weighted search provides a faster and more accurate method for validation of microarray probes.

A sample 25mer probe (Figure 1) is used with linear weighted and centre weighted searches against a SynaBASE of the full Human genome. The result of the linear weighted search (Figure 2) shows a contrast to the centre weighted search (Figure 3), in which there are more targets with mismatches biased towards the ends of the probe. Results of searches with BLASTN (Figure 4) and BLAT (Figure 5), using fixed k-mer seeds miss targets found by SynaHybridise. In terms of performance, both BLASTN and BLAT would require a cluster, while SynaHybridise runs on a single server to return the result within seconds. This clearly demonstrates the advantage of using an ungapped, non-heuristic weight matrix search approach as an improved alternative to verify the specificity and uniqueness of microarray probes.
To try SynaHybridize, please:
  1. Click here and then click on the “Access Now” button.
  2. Click “SynaHybridise”.
  3. Click “Test Sequence” (written in grey on the left hand side)
  4. Click ‘Copy’ to select the newsbyte test sequence.
  5. Select the ‘Linear Weighted’ search option and set ‘Maximum Mismatches’ to 4. Click ‘HYBRIDISE’ to begin your analysis.
  6. Go back to the query page, and now select the “Centre Weighted” search option and set the “End letter mismatch score” to -0.4, and set “Threshold” to -4. Click ‘HYBRIDISE’ to begin your analysis.

Figure 1:  Input page with the sample probe and parameters.

Figure 2:  Linear weighted search finds targets with mismatches at equal penalty settings.

Figure 3:  Centre weighted search finds targets with mismatches biased towards the ends of the probe.

Figure 4:  BLASTN searches are restricted by word size (word size 7, e-value 1000), and miss other potential cross talk targets that SynaHybridise identified.

Figure 5:  BLAT only reports one exact match.