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The sequence region to be used as the detection template must be considered carefully. Regions wit obvious secondary structure or long runs of the same nucleotide (e. g. with. non-coding regions) should be avoided. For detection of coding sequence specific to RNA targets it is advisable, where possible, to design the probe to span exon-exon boundaries. This prevents the detection of sequences from residual genomic DNA in the RNA prep. In circumstances were this not an option, the RNA sample should treated with DNAase prior to the RT step. This is an efficient approach and results in minimum loss of sample when carried out on a column-based purification system.
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