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Nucleic Acids Research, Vol 24, Issue 16 3278-3279, Copyright © 1996 by Oxford University Press

ARTICLES

An efficient method for generation and subcloning of tandemly repeated DNA sequences with defined length, orientation and spacing

SW Jiang, MA Trujillo and NL Eberhardt
Department of Medicine, Mayo Clinic, Rochester, MN 55905, USA.

Tandemly repeated DNA sequences generated from single synthetic oligonucleotide monomers are useful for many purposes. With conventional ligation procedures low yields and random orientation of oligomers makes cloning of defined repeated sequences difficult. We solved these problems using 2 bp overhangs to direct orientation and random incorporation of linkers containing restriction sites during ligation. Ligation products are amplified by PCR using the linker oligonucleotides as primers. Restriction digestion of the PCR products generate multimer distributions whose length is controlled by the monomer/linker ratio. The concatenated DNA fragments of defined length, orientation and spacing can be directly used for subcloning or other applications without further treatment.
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