This characteristic is exploited in TOPO-cloning. 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. 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. The linear vector DNA already has the topoisomerase enzyme covalently attached to both of its strands' free 3' ends.
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Cloning and DNA Markers
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