Running a combinatorial Mock, SLIC/Gibson/CPEC or Golden Gate DNA assembly design on the j5 server
Once we have created a collection object that is "j5 ready" (and a combinatorial design), we can use DeviceEditor to directly run a Combinatorial Mock, SLIC/Gibson/CPEC or Golden Gate DNA assembly design on the j5 server.
Here is a demonstration video that goes through this process (Run j5 Combinatorial Golden Gate assembly design.mov, right click this link to save the movie to disk for better viewing):
Here is the zip file (containing the resulting j5 output) resulting from the demonstration above (j5_njhillson_20120527030535.zip):
(Note: to be "j5 Ready" and a combinatorial design, each collection bin must contain at least one mapped part icon, and at least one bin must contain more than one mapped part icon).
If we click on the "Collection Info" tab in the right panel of the DeviceEditor display, we can see the collection's properties. In the "Collection Info" tab, the "j5 Ready" status will reflect a green "True" if the collection is ready for j5 DNA assembly design. If so, the "Combinatorial" status will reflect a blue "False" if there is only one part in each column or a purple "True" if there is at least two parts in one of the columns.
For more information on designing Combinatorial Golden Gate DNA assemblies with j5, refer to the "Combinatorial Golden-gate assembly" page in the j5 user's manual. Note, however, that the example provided in the j5 user's manual utilizes the j5 simplified web interface (and not DeviceEditor); as such, many of the steps described therein are analogous to, but distinct from, how they are accomplished here using DeviceEditor.
The video demonstration above closely mimics in intent the "Combinatorial Golden-gate assembly" example in the j5 user's manual (which utilizes the j5 simplified web interface). Steps 1-3 in the j5 user's manual are equivalent to creating part icons, and mapping sequences to them in DeviceEditor. Step 4 is equivalent to creating a collection object to contain the parts. Step 5 is equivalent to creating Eugene design specification rules for the parts. Step 6 is equivalent to the process described here, below. Step 7 is equivalent to controlling the j5 design parameters. Step 8 is equivalent to the process described here, below. Steps 9-11 remain the same whether using the j5 simplified web interface or DeviceEditor. The demonstration video above starts at a point where the equivalents to steps 1-5 and 7 have been completed.
With the "j5 Ready" status indicator a green "True", and the "Combinatorial" status indicator a purple "True", in the panel on the right side of the DeviceEditor display within the "Collection Info" tab, it is possible to use DeviceEditor to directly call the j5 server to design a Combinatorial Golden Gate DNA assembly protocol for the collection. Click the "j5" button at the top left of the DeviceEditor display. This will pop-up a new "j5 Controls" dialog box. Within this dialog box, click on the "Run j5 on Server" tab. Within this tab, you will have two options available for the "j5 Parameters File": either "Use lastest server version" (which will use your most recently updated set of j5 parameters currently stored on the j5 server) or "Generate file from parameters" (which will generate the j5 parameters file on-the-fly, based on how you set the j5 parameters for this design in DeviceEditor). You have three options available for the "Master Plasmids List", "Master Oligos List", and "Master Direct Syntheses List": either "Use lastest server version" (which will use your most recently updated corresponding master list stored on the j5 server), "Generate empty file" (which will generate the a new empty list file on-the-fly, using "j5_" as the initials and starting with number 1 when naming the plasmids, oligos, and direct syntheses), or "Choose File" (which will upload the corresponding master list file to the j5 server, only files with .csv extensions will be selectable). Note: the file names for these master list files (or the first named plasmid, oligo, or direct synthesis piece contained within them), determine how the resulting designed plasmids, oligos and direct syntheses will be named (and numbered); see, for instance, the "Master plasmids list file" description page in the j5 user's manual for more information. While the "Generate empty file" option is a nice way to start using DeviceEditor and j5 with minimal effort, it is recommended that you use the j5 feature described in the previous sentence so that your plasmids, oligos and direct syntheses will be named according to your initials and current numbering (or whatever your preference may be) instead of the default "j5_" and starting with number 1. Finally, there are two options available for "Assembly Method": "Combinatorial Mock", "Combinatorial SLIC/Gibson/CPEC" or "Combinatorial Golden Gate". For this example, we will be selecting "Combinatorial Golden Gate".
After you have set the "Design Assembly" options, click the "Run j5" button to initiate the design process on the j5 server. After doing this, you'll see the message "Your request has been sent to the server. Waiting for response...". For combinatorial assembly Golden Gate designs, the design process will take an amount of time commensurate with how many combinations you are asking j5 to design (the example above, with 8 distinct plasmids designed, takes a little less than a minute). Designing Golden Gate assembly designs for which it is especially challenging for j5 to find a set of compatible endonuclease overhangs may add substantially to the required run time. You'll likely eventually see the status message "Still waiting... This may be normal, especially for larger designs.". Once the design process has completed, the status message and progress bar will disappear, an "Update Part Preferred Overhangs" button and a "Download Results" button will appear, and the (sorted) names of the resulting designed plasmids will be listed in the "Plasmids" tab. Clicking on the hyperlinks provided for each of the resulting designed plasmids will open the corresponding sequences in VectorEditor for immediate visual assessment. Clicking on the "Download Results" button will pop-up a new "Select location for download" dialog box, which will allow you to select where you'd like to save the resulting zipped j5 output file.
For more information on what is contained within the zipped j5 design output file, refer to the "Combinatorial Golden-gate assembly" page and the "j5 output" section in the j5 user's manual.
After you have examined the j5 output and have decided that you are satisfied with the design, clicking the "Update Part Preferred Overhangs" button will add the j5-designed 5' and 3' preferred overhang positions (if any) to their respective parts in the design. For more information, refer to the "Target part order list file" page in the j5 user's manual.
It is also possible within the "Run j5 on server" dialog box tab to click the "Load Existing Assembly File" button, which will pop-up a new "Select file to upload" dialog box, prompting you to select a zipped j5 design output file. Doing so will populate the "Plasmids" tab will all of the plasmids designed within this zipped j5 output file.