Construction of larger-size sequencing templates from degraded DNA.

We describe a modified recombinant PCR procedure that allows the construction of longer fragments of DNA from shorter, overlapping PCR products. This method permits the construction of optimal-size templates for DNA sequence analysis (500-600 bp) from DNA that is too degraded for direct amplification of fragments of this size. As an example, the entire hypervariable region 1 of human mtDNA (418 bp) was reconstructed in one step from five overlapping amplification products (159, 126, 131, 131 and 93 bp) derived from a Paleolithic tooth specimen.