US 6,150,111 A · Methods and kits for recombining nucleic acids

Abstract

The invention includes a method for recombining and, optionally, cloning nucleic acids without the use of restriction enzymes or DNA ligase. The method involves recombining an insert and a recipient nucleic acid using custom designed complementary regions to anneal strands from the nucleic acids. The complementary regions on double stranded nucleic acids are exposed by limited digestion of the ends of the nucleic acids. After annealing the digested ends, single stranded gaps on the hybridized nucleic acid are closed to yield a double stranded nucleic acid which may, optionally, be cloned into a microorganism. The invention also includes kits for recombining and, optionally, cloning nucleic acids using the methods of the invention. The invention further includes recombinant nucleic acids prepared using the methods of the invention.

Claims17

Dependent claims

A method of recombining an insert nucleic acid and a vector nucleic acid, said method comprising

a) digesting the 5' terminus of an end of said insert nucleic acid to yield a 3' overhanging insert portion and a 5' digested insert terminus;
b) digesting the 5' terminus of an end of said vector nucleic acid to yield a 3' overhanging vector portion and a 5' digested vector terminus, wherein said 3' overhanging vector portion and said 3' overhanging insert portion are complementary;
and
c) thereafter annealing said 3' overhanging vector portion and said 3' overhanging insert portion to yield a double stranded complementary region, whereby said insert nucleic acid and said vector nucleic acid form a recombined nucleic acid, wherein both said insert nucleic acid and said vector nucleic acid are prepared by PCR and further wherein neither said insert nucleic acid nor said vector nucleic acid is prepared using a restriction endonuclease.

1 more element

Dependencies are inferred from the claim text.

Methods and kits for recombining nucleic acids

Life of this patent

Methods and kits for recombining nucleic acids

Life of this patent

Abstract

Claims17