Create oligo duplex and oligo conjugate entities

Modeling the connection of a biologic with small molecules creates both complexity and variability in a system of record. The oligo conjugate and oligo duplex tools in Benchling allow you to model oligo conjugates and oligo duplexes by bringing together oligo entities and molecule entities into a composite entity to visualize, validate, and design new bioconjugates, such as siRNA and ASOs, in a structurally accurate manner. 

If you are modeling oligo conjugates or oligo duplexes with the complex polymer schema, that means your team's admin has set up a single schema that defines oligo schema(s) as the backbone and other schemas like molecules as components. For more information on these schemas, see the Configure oligo conjugate and oligo duplex schemas article. Because oligo conjugate and oligo duplex schemas are composites of multiple Benchling schemas, it means that the individual component entities must be created and registered before you can create your oligo conjugate or oligo duplex entity. For more on how to create the component entities, see Creating and importing molecules.

Once the component entities are created, you can create your oligo conjugate entity. There are a few steps in that process, which at a high level include: 

1. Setting leaving groups on your molecule entities
2. Creating the oligo conjugate entity
3. Creating the attachments in the entity
4. The process can also include editing attachments as you iterate on your process 

Leaving groups on molecules 

Groups of atoms in a molecule entity can be set as attachment sites, or leaving groups, to model connections between backbone and conjugate entities. Setting a leaving group will subtract those atoms from the final structure and the final molecular weight of the conjugated molecule. Leaving groups will have assigned integer values as a means to identify those leaving groups when viewing the original molecule, and during the attachment process when forming an oligo duplex or conjugate.

Setting leaving groups on molecules is not a requirement for creating a complex polymer entity, instead you can draw the chemical attachment between the components. See section “Creating conjugates via drawn attachment module” for more information.

Note: Setting leaving groups does not affect the structure of the entities. The labels provide a technical pathway for groups to be used when designating connections between entities. 

Set a leaving group

1. Identify the molecule you wish to prepare to add to an oligo duplex or oligo conjugate entity and open its Chemical Structure tab
2. Select the Edit icon
3. Highlight a group of atoms to designate as leaving groups
4. Right click the highlighted atoms and select Atom Properties.
5. To assign the highlighted atoms as a specific leaving group with an assigned integer, click the lock icon to unlock the Map field and label the leaving group with an integer value that has not already been used
6. Accepting the changes by clicking ok
   • Atoms that are designated as part of the leaving group will be labeled with the Mapping number
7. Click Save to update the molecule entity 

This leaving group on this molecule can now be selected when you create an oligo conjugate or duplex.

Create the oligo conjugate or duplex entity

Individual complex polymer entities can be created via many methods in the platform. 

Create the entity with global create

When creating them through the Navigation bar pathways, you can create and designate attachments in the same process. 

1. Click the + on the left-side menu. Under complex polymer, select an oligo conjugate or oligo duplex schema
2. Enter the name and select a folder to store the complex polymer in
3. Define the conjugate
   • For an oligo conjugate: Select the component entities and backbone
   • For an oligo duplex: Select the forward and reverse strands, offset, and component entities

5. Optionally configure attachments, which can be set during creation or later
   • If you choose to set attachments after creation, you can use the drag and drop feature to set the attachment. This is explained "Edit existing attachments" section of this article
6. Click Create 

Select attachments during creation

After ensuring that the molecule has the proper leaving groups configured, attach the molecule to an oligo backbone. Optionally, multiple molecules can be attached to each other, or the backbone. 

1. Click the Configure attachments button
2. Select the leaving group on the component
   • If more than one component was defined, you can choose to conjugate to another component, or the oligo backbone
3. Set the index for the backbone. This will populate the available monomer for the conjugate to attach to
   • Note: the capping groups and selection group needs to be available in order to attach. If you would like to draw the attachment instead, proceed to “Creating conjugates via drawn attachment module”
   • For an oligo duplex, the index can be specified on either the forward or reverse strand
4. Click Attach to save the attachment.
5. If multiple molecules are being used, the system will automatically navigate to the next molecule to set attachments

6. Repeat steps 2-5 as necessary for additional components
7. After appropriate attachments have been set, click Create

Draw attachments during creation

For more custom chemical attachments, or for an alternative workflow, you can use Benchling’s MarvinJS modal to draw the attachment between the oligos and components.

1. Click the Configure attachments button
2. Select the Backbone entity you’d like to attach your first component to
   • If more than one component was defined, you can choose to conjugate to another component, or the oligo backbone
3. Set the index for the backbone. This will populate the available monomer for the conjugate to attach to
   • For an oligo duplex, the index can be specified on either the forward or reverse strand
4. Use the Marvin JS tool to remove necessary sections of your molecules, and draw their connections
   • If multiple molecules are being used, the system will automatically navigate to the next molecule to set attachments
5. Click Attach and Create to create your new entity

Edit attachments of oligo conjugates and duplexes

If attachments need to be made or edited after an entity has been created, this can be done from the polymer components tab on an oligo conjugate or duplex entity.

Edit existing attachments

If you have created an entity without configuring the connection, you can use the drag and drop interface to set the connection, otherwise use the clicks described below to make any edits. 

1. Navigate to the polymer map tab
2. Click the Edit icon in the bottom right corner of the polymer map to edit the conjugate
3. To maintain the index that the molecule is attached to, double-click the line representing the attachment that requires updates
   • Adjust the leaving group, if applicable
   • Click Attach
   • Save the new structure by selecting the Pencil icon again

To change the position that the molecule is attached to

1. Navigate to the polymer map tab
2. Click the Edit icon
3. Click the line connecting the molecule to the backbone, and select Delete on your keyboard
4. Move the molecule to a new position in the polymer map
5. Set the attachment point
6. Select Attach
7. Save the new structure by selecting the Pencil icon again

Add new molecule entities 

1. Open the polymer components tab
2. Select the pencil icon next to the “components” header to add additional molecule/oligo entities to attach to the backbone(s)
   • Note: the components section will list any component that has been added to the conjugate, but not attached yet.

3. Set attachments between the monomers of the backbone oligo and molecules in the attachment modal
   • R groups of the backbone are designated in the monomer library. These capping groups are subtracted from the final structure, the same way as in a nucleotide sequence.