Is it possible to use nested PCR techniques?
Nested PCR can be used to increase sensitivity, however the reproducibility of relative quantification will be lower. The labelled SALSA forward PCR primer starts at the 5’ end of the ligation products. Replacement of all short synthetic probe oligonucleotides by longer ones will provide the possibility to design a nested PCR primer. The unlabeled SALSA reverse PCR primer binds 14 nucleotides from the 3’ end of the ligation products. Nested PCR techniques will not remove most ligation-dependent background. If the sensitivity of the current MLPA protocol is insufficient for your application, please contact MRC-Holland.
