The Sample Multiplexing tool performs Sample Multiplexing on the aliquots of selected plates. This tool is used previous to the DNA sequencing process. Without "barcoding" the DNA elements, it finds the best way of pooling these aliquots together in order to reduce sequencing costs as much as possible.
Output: Pooling instructions
- (required) The plate(s) must already exist in inventory (Inventory > Plates)
- If the plate does not exist yet, use go to Inventory > Plates > ‘New Plate’ or ‘Upload’ to create a new plate.
- (required) Selected plates must have wet aliquots.
- If the aliquots are dry, use another tool (like the Normalization Planning tool to hydrate the aliquots).
You can find the ‘Sample Multiplexing’ tool under Tools > Tool Library. Click ‘Launch Tool’ to start using the tool.
The tool will walk you through a few steps to select samples and configure parameters before running. You need to complete each step before proceeding to the next. After completing a step, the completed step mark (circle) will turn green with a checkmark. You can always use the ‘previous’ button to go back to any previous step.
1. Select Samples: Click ‘Select Plates’ to select one or more plates (containing the samples that you want to multiplex) from inventory by checking the box(es) next to the desired plate(s). Click ‘Select Plates’ to confirm your selection.
After selecting the desired plate(s), you can view the table of selected plates. To change the selections, click ‘Change Plates’, or click on the red trash can icon above the table to remove all selected plates from the list. To remove a single plate from the list, click on the red trash can icon at the right of that plate’s row. Click on the eye icon at the left of a plate’s row in order to see the plate map and listed contents.
You will also see a table with all the samples to be multiplexed, contained in the selected plates. If you wish to ignore any sample(s), you can check the 'Ignore' checkbox to the right of the corresponding sample. When satisfied with the selections, click ‘Next’ to go to the next step.
2. Configuration & Submission: In this step, you will be able to configure the parameters of the sample multiplexing tool. These parameters define how the sample multiplexing algorithm is executed.
Here are descriptions about each one of these parameters. These descriptions are also visible to the user when clicking on the tooltips next to each parameter name.
- Maximum Alignment Overlap: A pair of samples/aliquots is going to be restricted from being pooled together if the length (in base pairs) of any of the found DNA regions between them exceeds this parameter.
- Maximum Tuple Size: Maximum number of samples/aliquots that are allowed to be pooled together.
- Exclusive Annotations: A list of annotations from which to compute the overlaps. The user may want to compute overlaps only on certain regions of the reference sequence for the 'overlap computation step'.
- Excluded Annotations: A list of annotations to exclude. The user may want to exclude certain DNA regions of the reference sequence from the 'overlap computation step'.
After specifying these parameters and naming the data table that will be generated, click on the ‘Submit’ button. You will then be notified that the Sample Multiplexing tool has completed and there is a link to the new data table that stores the output of the algorithm.
If you click on the link to 'Created 1 Table (pooling tool) you will be brought to a data table of type 'Pooling Schema'. Each element in the table specifies a sample/aliquot, and it shows its corresponding 'Pooled Sample Name'. The tool will have pooled one or more sample/aliquots into a specific 'Pooled Sample'.