Find Oligonucleotides.

The database constructed, and target taxon identified, the next step is to identify all possible oligonucleotides that could describe the selected group.  At this stage we are only interested in identifying oligonucleotides shared by as many of the sequences in the target group as possible.  Later, these potentially useful oligonucleotides will be screened for their ability to exclude non-target records. 

1) Select 'Find Oligonucleotides' from the main window.  Records within the target group () are displayed in the dialog box table:



2) Using the check boxes in the right-most table column , select representatives of the target group from which to generate/design the oligonucleotides. 



If the selected target group is small (e.g. < 10) selecting all will probably be the best strategy.  However, with larger target groups it may be more productive to select only a proportion (say, no more than 15), as too large a selection may resulting in the program missing oligonucleotides, which whilst  able to describe every single target record, may not be effective in descringuishing non-targets fronm targets (choosing ~15 ensures a compromise between finding the best oligonucleotide which describes the target group, and one which exclused the most non-target records).
At least one target record must be selected to design from, for the program to continue.

3) Use the 'Required length' spinner box to select the required oligo length (between 3 and 100 nucleotides).  Use the 'Maximum degenerate bases' spinner box to select the maximum number of acceptable degenerate bases.


4) Once you are happy with your selection, click the ' Find Oligonucleotides' button.  The program will then collect together the selected sequences, align the sequences (if necessary) and generate a consensus sequence from which the oligonucleotides are then generated.   Once complete, the total number of putative oligos identified is quoted along with a breakdown of how many have each sort of degeneracy.


5) If you are happy with the number of oligos generated, click ' Accept' to proceed to the next stage.  ' Reset' buttons clears all selections. ' Cancel' returns to the main panel without accepting oligos.  Tip: the speed of the next stage of the program is partially dependent on the number of putative oligos identifed.  Thus, if a very large number of oligos are identifed at this stage (e.g. >700) it might be better to amend parameter choices (i.e., reduce maximum degeneracy; select more target records to design from) to reduce this figure, to a more managable level.