Polimerase Chain Reactions (PCRs) are on the list of the most commonly used molecular biology techniques and can be part of the daily workday tasks in the lab. PCRs can increase the volume of existing aliquots or create new materials to add to the Lab Inventory.

The primer design is a very important step in this process. Your primers need to be carefully designed to be specific to amplify only your desired product (or to have the functionality you want, like creating an overhang, mutation, etc.). You can either manually create your primers or do it automatically with TeselaGen.

For this example, let's amplify a dCas9 enzyme from a plasmid in our DNA sequences library (you can follow this tutorial with any other genetic piece you want). First, open your DNA sequence and select the region you want to amplify:

Once your region is highlighted on the Open Vector Editor, right-click on it and select Create > Create PCR primers for this region to automatically create them.

This will open a pop-up window where you can select the strand (positive or negative for primers forward or reverse), tag your primers, and indicate your desired min and max values for melting temperature (Tm) and size.

Once you create your primers, you need to create a DNA Assembly Report to simulate the PCR. A pop-up window will appear, where you will find the "Submit for Assembly" section. Now, let's check our Assembly Report by clicking on the notification indicating that it's done (or going to the reports library).

Here, you need to scroll down until you find the "Assembly Oligos" section. In this section, click on the option "Save to Oligo library" if you want to save your primers.

Now, let's see how we can use the primers to obtain the expected PCR result.

In your DNA Assembly Report, you will see the section "PCRs". This contains the pieces obtained if you carry out a PCR with the primers you designed. Remember that, in this example, we created primers to amplify a dCas9 enzyme.

Let's click on the PCR_0001 and see what information is in it:

Here, we can see that our generated PCR result is linked to our DNA Assembly Report, but how can we check if we obtained the desired piece? One quick way is checking the length of the piece. In this example, our obtained size is 4104 bp, which is the same as the dCas9 size of the original plasmid that we wanted to amplify. However, we can also open the Open Vector Editor to see the expected sequence:

In this sequence, you can see how the generated oligos bind to the start and end of our designed sequence, successfully amplifying the complete CDS.

You need to notice that this is a PCR simulation, not a real experiment. Then, TeselaGen's system understands that you are checking how your primers would perform but not doing it in the lab. This means that your generated primers and PCR results don't exist in your Inventory, and none of the actions described previously would have any effects in your LIMS.

If what you want is to actually carry out a PCR in your lab with primers and DNA pieces that physically exist, then you need to use the Run PCR Tool, which needs you to have all the required materials in your inventory, and will generate a worklist to update or create new plate materials with your PCR results.