"Golden Gate" is a type of DNA Assembly that uses type IIS Restriction Enzymes to create a scarless design. TeselaGen's platform allows you to create a combinatorial design using this assembly method. In this tutorial, we will build a small combinatorial DNA library of 8 constructs. Next, let's see how to proceed with it:

For this design, we want to divide each construct into the following components:

Notice that in some components we are adding more than 1 variant. This allows you to create a combinatorial design.

We will start by creating a new design called Small DNA Library. To do this, go to the "Designs" library on the Molecular Biology Toolkit. Once you're there, click on "New Design" and select the "Blank Design" option.

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We are saving the new design as "Small DNA Library", and selecting "combinatorial" as the layout type. Now, click on "Save".

After saving the design, the design editor will look like the figure below. Notice that there are "plus (+)" symbols next to the icons to add a new bin or row. The design information is accessible on the right-side panel.

Details of each bin (or component) can be viewed or modified by clicking the bin icon on the inspector panel on the right. Since each construct has 6 components, let’s add 4 more bins to the current design. In addition, the 2nd, 3rd, and 5th parts each have 2 variants, therefore, we want to add one more row to the current design. Doing so, our combinatorial design now looks like this: 

TeselaGen's design editor has a variety of symbols (SBOL icons) representing different genetic elements that could accommodate the needs of visualizing complex DNA constructs. To edit a bin, click on it; this will display the right side panel, where you can change the icon, name, and other settings.

In this particular design, the 2nd, 3rd, and 4th components are coding sequences; hence, it makes sense to use a “CDS” (coding sequence) icon for these components. For the last two components, we can use a “Protein Stability Element” icon. After changing icons and names, the sketch of our design now looks like the figure below:

Now, let's assign the actual DNA parts. Do this by double-clicking a cell you want to assign a DNA part to, and a pop-up window will appear for you to select from your DNA Parts Library.

For the components with variants (e.g. 2nd, 3rd, and 5th parts), select all the parts needed on the same window. Following these rules to assign DNA parts to all the cells in the design, the “Submit for Assembly” button turns green and becomes enabled. The final design with all parts should look like this:

Now, here is a trick. It turns out that these little degradation tags are pretty short. We need to do some PCR to pull some of our desired DNA out of their host vectors, so why not embed these little guys into the forward and reverse primers? We can tell the assembler to do this by assigning a "Forced Assembly Strategy".

You can select the forced assembly strategy options for any part by selecting the part of interest, then select the part icon on the inspector panel on the right to view the part’s details. For our example, let’s click on the "ssrA_tag_3prime" cell, then pick "Embed in primer forward" from the "Forced Assembly Strategy" dropdown menu.

Alternatively, you can right-click the part, and choose a strategy from the “Forced Assembly Strategy” menu. For the two DNA part options in the ssrA_5primeTag column, pick "Embed in primer reverse".

The design editor uses colored markers to indicate different assembly strategies (purple and green in this case, shown in small squares on the top left side of each part). These colors add a layer of information to the visualization of the design without overwhelming the users.

We can add one more feature to this design. It turns out that some of these parts are contiguous in their hosts, and TeselaGen's assembler is smart enough not to break everything apart, just to put it all back together again. However, the assembler will do a cost tradeoff between direct synthesis and PCR based on the cost of synthesis. To tell the assembler not to consider direct synthesis, you can impose a "Direct Synthesis Firewall" to a column. To do this, select a bin and click on the “Direct Synthesis Firewall” from the right side panel.

In our example, we will do this for columns 2 and 5. Our final design looks like this:

Finally, you can modify the assembly method by selecting it from the drop-down menu in the “Assembly Reaction Details” option on the side panel.

That is about as fancy as we need to get for this design. If you navigate to the green "Submit for Assembly" button at the upper right corner of the design editor,  you can run the assembler and build your library.