NEBuilder Assembly Tool can be used to design primers for your NEBuilder HiFi or Gibson Assembly reactions based on entered fragment sequences and the polymerase being used for amplification. This video will highlight some useful updates and the main differences between the two versions.
NEBuilder Assembly Tool can be used to design primers for your NEBuilder Hi-Fi or Gibson assembly reactions, based on entered fragment sequences and the polymerase being used for amplification. This video will highlight some useful updates and the main differences between the two versions.
Vectors Treated as Fragments. NEBuilder version 1 was designed primarily to assist beginners with cloning an insert into a vector. The primary fragment view emphasized this by showing a split vector flanking the inserted fragment. Behind the scenes, NEBuilder version 1 split the vector sequence into two fragments. This also affected the starting point of the assembled sequence. In NEBuilder version 2, vectors are treated similarly to any other fragment, with minor exceptions. This is reflected in the display, as the vector is now indistinguishable from insert fragments. The sequence of the vector, as with any fragment, is now kept intact rather than being split.
View of 6-Frame Translation. Another major change is how the 6-frame translation at a junction is viewed. In version 1, this view was accessed from the list of required primers. In version 2, junction details are viewed by selecting fragment. In addition to the 6-frame translation, the new detailed view displays the overlaps that were used to create the assembled junction.
Adding Custom Spacers. NEBuilder version 2 changes the point of which custom spacers between fragments can be added. In version 1, the spacers could be entered during the initial fragment entry, or by editing an insert fragment by double clicking on it. In version 2, spacers can only be added after fragments are entered by editing them, clicking on the pencil icon and scrolling down to the junction properties section.
Editing Method. Finally, editing methods are slightly different in NEBuilder version 2. Adding new fragments is basically unchanged. Click the appropriate button, add fragment versus new fragment. To delete in version 2, simply click on the X at the right end of the fragment and confirm. In version 1, the delete button brought up a dialog to choose which fragment to delete. Also, editing a fragment's properties is initiated by clicking the pencil icon in version 2. In version 1, a double click on the fragment opened the edit window, not compatible with mobile devices.
In addition to the changes already mentioned, NEBuilder version 2 includes a substantially updated user interface, better error handling, better assembly of the expected final sequence, and many new features based on user feedback, including:
Sequence files can be uploaded into the application. FASTA, Genbank and raw text files are supported.
Restriction digested fragments that are not classified as vectors may be used.
Restriction enzymes that cut up to 2 times are now displayed. The appropriate fragment can be selected when more than one fragment is produced by the digest.
Easier and more complete editing of fragment properties. Fragments can be fully re-specified without deleting and replacing.
Overhang distribution at junctions can be specified. Either all on one, or split between two fragments when possible.
Long sequences can be split into shorter overlapping fragments.
Projects can be saved to and reloaded from the user's computer.
Better detailed view of 6-frame translation and overlap between fragments.
Downloadable final assembly, component fragments, required oligos, and a PDF summary.
We hope these updates made designing primers for your DNA assemblies easier and more convenient. Please contact [email protected] with any questions.