The present invention provides compositions, methods, and kits for covalently
linking nucleic acid molecules. The methods include a strand invasion step, and
the compositions and kits are useful for performing such methods. For example,
a method of covalently linking double stranded (ds) nucleic acid molecules can
include contacting a first ds nucleic acid molecule, which has a topoisomerase
linked to a 3 terminus of one end and has a single stranded 5 overhang
at the same end, with a second ds nucleic acid molecule having a blunt end, such
that the 5 overhang can hybridize to a complementary sequence of the blunt
end of the second nucleic acid molecule, and the topoisomerase can covalently link
the ds nucleic acid molecules. The methods are simpler and more efficient than
previous methods for covalently linking nucleic acid sequences, and the compositions
and kits facilitate practicing the methods, including methods of directionally
linking two or more ds nucleic acid molecules.
