14.9 TOPO® Cloning
TOPO Cloning lets you ligate a single fragment into a Vector within only 5 minutes using the natural activity of Topoisomerase I which recognizes a speciﬁc motif 5′ - (C/T)CCTT - 3′ on the DNA. TOPO is a registered trademark of Invitrogen Corporation.
The TOPO cloning function under the Cloning menu allows you to insert linear fragments into either linear TOPO vectors (when a TOPO-site is present at the extremities) or into circular TOPO vectors. You can select as many sequences at once as you like, they will be ligated into each other in a batch operation.
TOPO Cloning options dialog.
- Three diﬀerent options (TA- Blunt- or Directional cloning) are shown on the top. If Directional is selected the user can deﬁne an overlap sequence. If this ﬁeld is blank it has the same eﬀect as Blunt cloning.
- The ﬁeld below shows which of the selected sequences have been detected as vectors, all other sequences are inserts.
- If any complications occur, eg. when more than one TOPO site is detected or when a linear sequence with TOPO site is selected it will print a message in this box, also showing how the corresponding sequence is processed if the user clicks OK.
- The resulting sequences will be optionally saved in a sub-folder.