View restriction enzymes

Benchling's restriction enzyme feature allows users to create, manage, and utilize custom lists of restriction enzymes, as well as search and filter a comprehensive enzyme library. This feature streamlines molecular biology workflows, particularly for cloning and DNA manipulation, by enabling scientists to quickly find enzymes that meet specific criteria, such as cutting at particular sites or exhibiting high-fidelity characteristics. With these tools, scientists can efficiently identify and apply the right enzymes for their experiments and analyze their impact on DNA sequences.

Create a new enzyme list

You might want to create a new custom enzyme list when you need to group specific restriction enzymes for a particular project or experiment. The configured enzyme list can then be viewed on any sequence map.This allows you to quickly access and manage a curated set of enzymes relevant to your current work, rather than sifting through an exhaustive enzyme library. Creating a custom list also enables you to control who can view the list within your organization, facilitating collaboration and consistency across a team.

1. Click on your Avatar, then navigate to Enzyme Lists from the Molecular Biology Settings sub-menu

   

2. Use the dropdown list to change the owner of the list and control who can see it in the tenant. You will be able to choose either yourself or an organization you are part of.

3. Under New List, use the text boxes to give your enzyme list a name and optional description, then click Create Enzyme List

   

4. Add enzymes to your list by searching through the enzyme library to find the ones you’d like to add. Once you’ve selected all the enzymes you want to add to your list, click Save changes.

Edit existing enzyme lists

After creating enzyme lists, you can edit an existing enzyme list to update its name, description, or manage its collaborators. This allows you to refine the purpose or scope of the list, ensure its details are accurate, and control who has access to or can modify the list, which is particularly useful for team projects or evolving research needs. To edit an existing enzyme list, you want to be in the same Enzyme Lists page of the Molecular Biology Settings.

1. Choose the desired enzyme list from the dropdown under Open Existing Enzyme List
2. Click on the Settings icon to get to Enzyme List Settings 

3. In the resulting pop-up, make your edits to the enzyme list name and once done click Save

Search for enzymes by name or number of cut sites

You might want to search for enzymes by name or number of cut sites when you are performing a digest analysis on a DNA sequence. This allows you to quickly locate specific enzymes or filter for enzymes that cut a particular number of times, directly within the Digests panel. This functionality helps in efficiently identifying suitable restriction enzymes for your experimental design. In an open DNA sequence: 

1. Click on the Digests icon in the molecular biology toolbar

   

2. In the panel that opens, there is a Find Enzyme search box that allows you to search for enzymes by entering the name or number of cuts
3. For example, enter “EcoRI” to pull it up in the list
4. Hover your cursor over the enzyme to see information on its activity, and a list of possible cut sites on the open sequence, clicking on a specific site will jump to it on the sequence
5. Once you've decided on the enzyme(s) you want to use, click on it. The cut site on the map will be highlighted on the sequence map. If you wish to unselect the enzyme, click it again

If you are unable to see the cut site listed on the sequence ensure all enzyme cut sites are turned on for your sequence of interest

Filter by advanced parameters

You can also filter enzymes by where or how many times they cut on the sequence. For example, you can filter to see only enzymes that are single or double cutters.  

Note: these filters will control what enzymes are seen on the sequence map and not in the list.

After you've selected part of a sequence, open the Digests panel. Use the dropdown at the bottom of the restriction enzyme/digest panel to specify what enzymes you see based on a highlighted region in the sequence. You have the ability to select enzymes that are in the selection, outside of the selection, etc.

How to run virtual digests 

Benchling's virtual digest feature allows users to simulate restriction enzyme digests on DNA sequences and visualize the predicted gel electrophoresis results. This feature provides significant value by enabling scientists to design and optimize their cloning and DNA manipulation experiments computationally before performing them in the lab, saving time and reagents. Additionally allowing them to compare their gel results to the expected results from the virtual digest ensuring the correct products were formed. The virtual digest targets workflows involved in molecular cloning, plasmid construction, and DNA analysis. 

1. Click on Digests in the molecular biology toolbar
2. Use the Find Enzyme search box to search for enzymes by entering the name or number of cuts
   • Note: Follow the above section to create your own enzyme lists for your specific organization 

3. After you've selected your enzymes, click Run Digest
4. A Digest tab will open in the sequence that shows details about resulting fragment(s) and reaction conditions. If you want highlight a fragment on the sequence tab, click the corresponding row in the fragment table. 

5. The Virtual Digest tab allows you to see the expected gel product of your digest

7. If you'd like to be able to reopen your digest, return to the Digest tab and use the text box to give it a name. Then click Save and you will be able to open the digest from the Saved Digests section of the Digests panel.

8. If you follow the steps above and perform another digest on the same sequence then the products will appear in the same virtual digest tab. You can then reorder the lanes of your virtual digest by reordering your digest tabs on your sequence.

How to Create Custom Ladders

If the pre-defined ladders do not match the specific band sizes or markers you are using in your physical lab experiments, then you may want to create your own custom ladder for a virtual digest. This allows you to accurately compare your virtual digest results to your actual gel electrophoresis results by providing a customized reference that mirrors your experimental setup. To create a custom ladder: 

1. Click on your Avatar, then navigate to Ladders from the Molecular Biology Settings sub-menu,
2. From there, you can create new ladders with custom band sizes by typing the size of the band in base pairs in the text box. Those new ladders show up as a selectable ladder in your virtual digest module.

Frequently asked questions 

Q: When I click high-fidelity enzymes, why are other restriction enzymes marked unavailable?

A: When you click high-fidelity (HF) to search for HF enzymes, this masks enzymes that do not have a high-fidelity version. This does not mean that there are no cut sites for these enzymes, only that there isn't a HF version. 

To see all enzyme options, toggle between checking and unchecking high-fidelity.

Q: What is a double cutter in the Benchling context?

A: A double cutter refers to both:

• enzymes where a single enzyme always makes 2 cuts for one recognition site
• enzymes that cut twice or more in the same sequence (because there are two or more recognition sites)