In conclusion, a target with one to nine evenly distributed mismatches, preserving multiple contiguous matching stretches of at least 5 nt, has little reducing effect on hybridization with a 70-mer probe, while a target with >14 evenly distributed pm hybridizes inefficiently or not at all. Hybridization can be improved by utilizing a less stringent (lower) hybridization temperature. Furthermore, grouped mismatches tend to strengthen the hybridization compared to evenly distributed mismatches. Consequently, for hybridization between 70-mer strands with 10-20 mismatches, the distribution of the mismatching nt affects the hybridization more than the number of mismatches, indicating that the length and number of perfectly matching stretches are of greatest importance.