Prerequisites

Each sequence in the submitted alignment must have an end placed numerical value in the FASTA header, separated from the rest of the line with a blank space e.g.

• >MySequence1 1.2
ARNDCQEGHILKMFPSTWYV

Scientific numbering will also be accepted, e.g.

• >MySequence2 1e-3
ARNDCQEGHILKMFPSTWYV

There must be at least two sequence-variants submitted and at least two different sequence associated values.

Submission

Please note that only the 20 standard amino acid residues (ARNDCQEGHILKMFPSTWYV) will be considered, any other characters will simply be excluded from the evaluation. This also includes gaps, which are thusly neither evaluated.

The sequences can be input in the following two ways:

• Paste a number of sequences in FASTA format into the upper window of the main server page.

• Select a FASTA file on your local disk, either by typing the file name into the lower window or by browsing the disk.

Options

Modify the default settings to fit your preferences.

• Significance threshold
  Choose a significance threshold beyond which you consider a residue as significantly distributed. α = 0.05 means that there is a 5% or less chance that the identified residue is in fact not significantly associated with the data set phenotype (Type I error, false positive)

• Method for correction for multiple testing
  From the drop-down list you can choose among different methods for adjusting the computed p-values. You can also choose to not correct for multiple testing. Bonferroni single-step is more conservative than Holm step-down. Choosing 'no correction' increases the chance that the identified residue is in fact not significantly associated with the data set phenotype (Type I error, false positive). Briefly: A p-value of 0.05 obtained when performing 10 tests, has a Bonferroni single-step corrected p-value of: p-value_corrected = min[1, p-value_{non-corrected} x n_tests] = min[1, 0.05 x 10] = 0.5

• Choose sorting of numerical values
  Decreased sorting means that the highest value is considered the 'strongest' (e.g. quantitative detection via fluorescence), increased in turn means that the lowest value is considered as the 'strongest' (e.g. binding affinity)

• Sequence identifier for relative numbering
  Enter the sequence identifier of a reference sequence into the text field. This sequence will be identified in the sequence set and all sequence positions in the heatmap and logo-output will be numbered relative to the reference sequence. If any gaps are present in the reference sequence, the gap positions will be numbered negatively, i.e. p_-1 is the first gap, p_-2 is the second and so on. Any non-gapped positions will be numbered sequentially p₁, p₂ ... p_n

• Type of logo
  Full logo: Include ALL residues at ALL positions.
  Significant positions: Include ALL residues at positions where at least ONE amino acid residue was identified as significantly associated with the data set phenotype.
  Significant residues: Include ONLY residues identified as significantly associated with the data set phenotype.

Submit the job

Click on the "Submit" button. The status of your job (either 'queued' or 'running') will be displayed and constantly updated until it terminates and the server output appears in the browser window.

At any time during the wait you may enter your e-mail address and simply leave the window. Your job will continue; you will be notified by e-mail when it has terminated. The e-mail message will contain the URL under which the results are stored; they will remain on the server for 24 hours for you to collect them.