Sense or Antisense? When designing a transcription template, it must be decided whether sense or antisense transcripts are needed. If the RNA is to be used as a probe for hybridization to messenger RNA (e.g. Northern blots, in situ hybridizations, and nuclease protection assays), complementary antisense transcripts are required. In contrast, sense strand transcripts are used when performing expression, structural or functional studies or when constructing a standard curve for RNA quantitation using an artificial sense strand RNA.
The +1 G of the RNA polymerase promoter sequence in the DNA template is the first base incorporated into the transcription product. To make sense RNA, the 5' end of the coding strand must be adjacent to or just downstream of, the +1 G of the promoter. For antisense RNA to be transcribed the 5' end of the noncoding strand must be adjacent to the +1 G. If the insert is in a vector, the vector should be linearized downstream from the promoter and the inserted sequence to be transcribed.
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