Processing Tracefiles Using the High Throughput View

To import more than 96 samples, it is necessary to use the Phineus High Throughput interface.

To do so you will first need to generate a list of files to be processed.

Select 'Generate a List of Tracefiles for Processing' from the 'Tools' menu.

In the new Window, select the tracefile directory that you wish to import by clicking 'Select Tracefile Directory' and navigating to the relevent directory using the file browser.

When you have the directory containing your files (Note: all tracefiles should be in the same directory), select open.

Phineus will then generate a table with the names of all the files in the selected directory.

Note: Phineus will not check to ensure that the file is an ABI tracefile - if there are non ABI files in the folder, you should remove them from the list, by highlighting them on the list, and selecting 'Remove Selected Files' located second from the bottom of buttons situated next to the table.

Once you have all of your files imported, you will need to supply the information describing the samples for Phineus as follows -

Firstly, select the project which the data will be associated with by selecting the project from the drop down list,

and then select 'apply to all':

You will notice that the table has now changed and all of the samples will show that their project is 'S. pneu MLST'

The dropdown menu for 'genes' will also now show the genes associated with that project.

If you have multiple different genes amongst your samples, then highlight the relevent samples for each gene in turn

and select the appropriate gene from the dropdown list headed 'Select Gene'

and then select 'Apply to Selected'.

Repeat this for each gene in your sample.
If all of the samples correspond to the same gene, then you can use 'Apply To All' insted of this method.

Next, you should supply the read direction of each sample. To do so, select those reads that are in the forward direction, and select the appropriate radiobutton

Now click 'Apply to selected'.
Repeat this process for the reverse direction.

Now you must input the sample names for which there are two possible methods.

1) If you have multiple samples from the same strain/isolate then it is reccomended that you select all of the isolates with the same name, and then write this name into the 'Strain' box:

and then select 'Apply To Selected' to avoid having to type the name out multiple times.

2) If the same isolate name only appears one or two times in the samples selected, then you can input this information directly into the table.

To do this, select the cell that you wish to change, and hit the return key, you can then edit the contents of the cell, and pressing return again will stop editing the contents of the cell.
If you then press hit return a third time, Phineus will move the cell selection down to the next row, allowing you to iterate through all of the sample names in the table, without having to use your mouse.

A similar method can be used to add and change other data in the table, such as the well name/id for each sample.

Once you have completed this process, you are ready to save the file. To do this, you should first select 'Output Names' and then navigate the file browser to where the tracefiles are located. Then choose an appropriate name for the tracefile name list, and select 'Save'.

You are now ready to Process the tracefiles.

To begin processing the tracefiles select 'Process Tracefiles HT Interface' from the 'Phineus' menu.

This will open up a new window showing the Phineus HT Data Importer

First, press 'Select Sample List File'. This will open a file browser, which you should use to navigate to the directory containing your tracefiles. Once in that directory, you should select the list file you created previously, and click 'open'.

You should then select the radiobutton marked 'Phineus Naming Scheme' before choosing the project which corresponds to the tracefiles to be processed.

Once you have done this, you can press 'Import'.

You should check that this list is correct, before selecting

This will then present you with a new page

You are now ready to start the automated processing of your tracefiles.

To do so, select

Your computer will then process the tracefiles, updating you on its progress as major components of the pipeline are completed.

Once this is complete, Phineus will take you to the Assembly screen, where you can view and edit your tracefiles.

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