How can I insert a restriction fragment generated by partial digestion into a circular vector?
A restriction fragment can be inserted at a restriction site in the vector, or it can replace a restriction fragment in the vector.
Display Sites on the Fragment File
Open the fragment file and use Enzymes → Choose Enzymes to display the site that will be used to generate the partial digestion fragment. See this article for details on how to choose enzymes – Choose Enzymes Manually.
In this example, a blunt partial RsaI fragment will be ligated into the vector-based SmaI site.
Start the Restriction Cloning Tool
View the vector file, then click Actions → Restriction and Insertion Cloning → Insert Fragment.
Set the Vector
In the Vector tab, use the dropdown to browse and choose the vector for cloning.
Select a restriction site or a pair of sites for insertion of the insert.
Define the Source Sequence
Switch to the Fragment tab, click the dropdown to set the "Source of Fragment" that contains the insert.
If required, click on the "Show Enzymes" button on the side toolbar to display the restriction enzyme site of interest. To learn more about displaying specific enzymes on a sequence, see the guide chapter on Enzymes.
Specify the enzyme used for digestion in the right panel. Initially a list of all fragments generated bu full digestion is shown.
Define the Partial Restriction Fragment
Click the dropdown and choose "Cut partially" and a list of all possible partial fragments will be displayed.
In the Map view select the partial fragment of interest.
Alternatively, in Map or Sequence view, click on the first site, then shift-click on the second site to select the partial digestion fragment of interest.
If there is an intervening site within the selected range then a list of all partial digestion fragments will be shown in the side panel. The selected partial fragment will be highlighted in the list.
Create the Product
Switch to the Product tab to preview the ligation of the partial insert into the vector.
Enter an appropriate name for the new product sequence.
Click Clone to create the new product sequence.
