1. Enzymatic Method for the Synthesis of Long DNA Sequences with Multiple Repeat Units
- Author
-
Bernard A. Connolly, Colette J. Whitfield, Andrew R. Pike, Eimer Tuite, and Andrew T. Turley
- Subjects
biology ,Base pair ,Chemistry ,DNA polymerase ,Stereochemistry ,General Chemistry ,General Medicine ,DNA ,Molecular biology ,Polymerase Chain Reaction ,Catalysis ,DNA sequencing ,law.invention ,chemistry.chemical_compound ,Polynucleotide ,Duplex (building) ,law ,biology.protein ,Nanotechnology ,Polymerase chain reaction ,Repeat unit - Abstract
A polymerase chain reaction (PCR) derived method for preparing long DNA, consisting of multiple repeat units of one to ten base pairs, is described. Two seeding oligodeoxynucleotides, so-called oligoseeds, which encode the repeat unit and produce a duplex with 5'-overhangs, are extended using a thermostable archaeal DNA polymerase. Multiple rounds of heat-cool extension cycles, akin to PCR, rapidly elongate the oligoseed. Twenty cycles produced long DNA with uniformly repeating sequences to over 20 kilobases (kb) in length. The polynucleotides prepared include [A]n /[T]n , [AG]n /[TC]n , [A2 G]n /[T2 C]n , [A3 G]n /[T3 C]n , [A4 G]n /[T4 C]n , [A9 G]n /[T9 C]n , [GATC]n /[CTAG]n , and [ACTGATCAGC]n /[TGACTAGTCG]n , indicating that the method is extremely flexible with regard to the repeat length and base sequence of the initial oligoseeds. DNA of this length (20 kb≈7 μm) with strictly defined base reiterations should find use in nanomaterial applications.
- Published
- 2015