Principle
The technique utilises the inherent biological activity of DNA topoisomerase I. The biological role of topoisomerase is to cleave and rejoin supercoiled DNA ends to facilitate replication. Vaccinia virus topoisomerase I specifically recognises DNA sequence 5ยด-(C/T)CCTT-3'. During replication, the enzyme digests DNA specifically at this sequence, unwinds the DNA and re-ligates it again at the 3' phosphate group of the thymidine base.
Read more about this topic: TOPO Cloning
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“I sincerely believe ... that banking establishments are more dangerous than standing armies, and that the principle of spending money to be paid by posterity, under the name of funding, is but swindling futurity on a large scale.”
—Thomas Jefferson (1743–1826)
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“Experimental work provides the strongest evidence for scientific realism. This is not because we test hypotheses about entities. It is because entities that in principle cannot be ‘observed’ are manipulated to produce a new phenomena
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—Ian Hacking (b. 1936)