Topo (cloning technology)
From Wikipedia, the free encyclopedia
 |
This article is uncategorized.
Please help improve this article by adding it to one or more categories, so it may be associated with related articles (how?). Please remove this tag after categorizing, but not before. |
The key to TOPO® Cloning is the enzyme DNA topoisomerase I, which functions as both a restriction enzyme and a ligase. Its biological role is to cleave and rejoin DNA during replication. To harness the religating activity of topoisomerase, vectors are provided linearized with topoisomerase I covalently bound to each 3' phosphate.
Vaccinia virus topoisomerase I specifically recognizes the pentameric sequence 5´-(C/T)CCTT-3´ and forms a covalent bond with the phosphate group of the 3´ thymidine. It cleaves one DNA strand, enabling the DNA to unwind. The enzyme then religates the ends of the cleaved strand and releases itself from the DNA.
