No abstract

Transposon Tn1O inserts preferentially at particular insertion "hot spots" that share a symmetrical 6-base-pair consensus sequence: 5' GCTNAGC 3'. The protein that recognizes this sequence is not known but is likely to be the TnlO-encoded transposase protein. We present evidence that the 5-methyl groups of the two thymines in this sequence are essential for efficient transposon insertion; in their absence the sequence is still recognized, but at lower efficiency. We have reached this conclusion by examination of a specific hot spot whose sequence is 5' GCCAGGC 3'. The innermost cytosines ofthis sequence happen to be substrates for methylation at their 5 positions by the bacterial dcm-encoded methylase. We find that TnWO transposes into this site 15 times more frequently in a Dcm' host than in a Dcm-host; in the Dcm-host, insertions still occur, but at a low frequency. Thus, at this site, the absence of pyrimidine 5-methyl groups at the third positions of the consensus sequence is sufficient to convert a strong insertion hot spot into a weaker but still recognizable hot spot. This observation supports the general proposition, suggested previously by comparisons among consensus sequences, that the presence or absence of these 5-methyl groups is one major feature that can make the difference between a strong and a weak TnlO insertion hot spot.Transposon TnJO can insert into many different sites in the bacterial genome. However, it inserts preferentially into particular specific DNA sites or "hot spots" (1). DNA sequencing of many different TnJO insertion hot spots has revealed the presence of a symmetrical 6-base-pair (bp) consensus sequence that plays a major role in determining TnWO insertion-site selection (2). This sequence is necessary but not sufficient to determine insertion specificity; other base pairs in the immediate vicinity and/or long-range structural features of the DNA also influence the distribution of insertions among different sites (2).Insertion of TnJO always results in the direct duplication of a 9-bp target-site sequence and integration of the transposon in between the duplicated copies ( Fig. 1; refs. 3-5). The existence of such "9-bp repeats" is interpreted to mean that the target DNA duplex is cleaved by a pair of appropriately spaced single-strand nicks during insertion (3, 6, 7). The symmetrical 6-bp TnWO insertion hot-spot consensus sequence is itself symmetrically disposed within the 9-bp repeat sequence in the target DNA. As shown in Fig. 1 TnJO results in duplication of a 9-bp target DNA sequence; the element is inserted between the duplicated copies. Transposition is presumed to involve staggered single-strand cleavages of the target DNA; the actual strand polarity of these cleavages is not known. The TnlO insertion-specificity consensus sequence is symmetrically located within the 9-bp target sequence.TnJO-encoded transposase protein, but no direct evidence to this effect is yet available.The consensus TnlO insertion-specificity sequence is 5' GCTNAGC 3' (where N represents...

Frequency of skin punctures is influenced by gestational age and postnatal age. Skin punctures were not an independent risk factor for sepsis.