Get started
Prepare PCR mastermixes By Volume Order
In this example, we will prepare three different mixtures by volume order, i.e., by adding first the liquid which is required in the highest amount and subsequently until the liquid which is required in the smallest amount. All mixtures will contain Water, buffer, MgCl2, DNA Polymerase and two primers, which vary for each mixture (1 to 3).
In the workflow builder, define your mixture components using the Define Liquids and Plates element. For more information on how to use the Define Liquids and Plates, click here.
Add one element to your canvas: Make Mixtures.
Connect the Liquids parameter from the Define Liquids and Plates element to the Liquids To Make Mixtures input parameter of the Make Mixtures element.
Click on the Make Mixtures element to open the element parameter window. Here you can specify your mixtures. First, specify
Parameters under Mixing Mode.
Now specify the composition of each mixture. To do so, give each mixture a name and add the names of the liquids defined in step 1. In this example, we will specify the volume of each component we want in each individual PCR reaction. We will then scale up the volume using a multiplier.
Note: because we are making the mixtures by Volume Order, the order in which each liquid is defined here does not matter.
Then specify which liquid should be used as a diluent. The diluent will only be used if the mixture composition is such that you need to top up with some volume to make the desired final volume or concentration. This can be specified globally for all mixtures or individually for each one.
Then, we specify the target volume for each mixture (which in this case is the volume of mastermix we would like to add to each PCR reaction).
Note: The sum of the volumes specified in step 7 is 16.5 ul and water is not defined. Considering that the Final Target Volume is set to 18 ul, 1.5 ul of water will be added to top up the difference.
Select Volumes (high to low) under Addition Mode.
Since we want to increase the target volume of the mixtures, but add their components in the same ratio, we set the Volume Multiplier input to specify the factor by which we want to increase its target volume = 100.
Finally, specify the number of replicates and how the mixtures should be laid on a plate. Select Let Synthace Optimise from the dropdown menu for Layout and Replicate Grouping.
Define a Plate Set Name. This can be any alphanumeric name such as Mixtures.
Select a Plate Type from your plate library. Take care to ensure the plate type you select can accommodate the final volume for your mixtures.
Simulate the workflow
Check that the device that you selected can follow the instructions that you prepared. To learn how, click here.
Preview the execution
After you simulate the workflow, click View Simulation to open the simulation details.
Open the Preview tab, then click through the steps to check that the instructions that Synthace has generated are correct.
You can see that mixtures are prepared, and the individual liquids are added in decreasing volume order, taking into account the volume multiplier:
1000 ul of Buffer
200 ul of Primer F1
200 ul of Primer R1
200 ul of MgCl2
150 ul of Water
50 ul of DNA polymerase
You can also easily inspect the composition of each mixture by selecting the corresponding well in the Setup view.
Check your work:
To see what your finished workflow to this tutorial should look like simply navigate to the Tutorials and search for Tutorial: 5. Make Mixtures.
Challenges
In the example above, the composition of each mixture was defined by the volume of each of its components. A mixture can also be specified by a desired target concentration of the components, or a mixture of volumes and target concentrations.
You will need to specify the source concentration of the source liquids in the Define Liquids and Plates elements. For tutorials on how to use this element, click here.