Construction of DNA recognition sites active in Haemophilus transformation.

Abstract

Competent H. influenzae cells recognize and preferentially take up Haemophilus DNA during genetic transformation. This preferential uptake is correlated with the presence on incoming DNA of an 11 base pair (bp) sequence, 5''-A-A-G-T-G-C-G-G-T-C-A-3''. To prove that this sequence is the recognition site that identifies Haemophilus DNA to the competent cell, a series of plasmids, each of which contains the 11 bp sequence was constructed. The ability of fragments from these plasmids to compete with cloned Haemophilus DNA fragments that naturally contain the 11 bp sequence was tested using 2 different assay systems. The addition of the 11 bp sequence to a DNA fragment is necessary and sufficient for preferential uptake of that fragment. Plasmid DNA containing this sequence may vary as much as 48-fold in uptake activity, and this variation correlates with the A + T-richness of the DNA flanking the 11 bp sequence.