Fig. 4

Flowcharts for molecular diagnosis of SRS (a) and BWS (b) syndromes. a. The 11p15.5 methylation test targeted to H19/IGF2:IG-DMR is the first assay and is currently performed by MS-MLPA or pyrosequencing or Southern blot. In cases with borderline methylation values, two or more combined techniques are useful and if applicable multiple tissues should be tested. Evidence of LoM with normal CNVs confirms the epigenetic defect, while LoM with altered CNVs suggests a microdeletion/duplication. These cases need to be defined by karyotyping, FISH, or array-based genomic methods which should be extended to probands’ parents in order to define the recurrence risk. Further proceeding in cases with the epigenetic defect is represented by the MLID test. Cases with normal 11p15.5 methylation levels are tested by chromosome 7 MS-MLPA and/or microsatellite analysis to evidence maternal upd(7)mat and possible epimutations. The negative cases are either processed by array-based genomic methods to detect upd at multiple chromosomal regions or sequenced for CDKN1C and IGF2 or addressed to differential diagnoses. b The 11p15.5 methylation test targeted to both H19/IGF2:IG-DMR and KCNQ1OT1:TSS-DMR is the preliminary assay, and alternative options are recommended according to altered (positive) or borderline results. Microsatellite analysis defines upd, while aberrant CNVs raise the suspicion of unbalanced translocation/duplication, which should be verified by karyotyping/FISH/array-based methods and, if confirmed, recommend parental analysis. Further proceedings are represented by MLID test in case of epigenetic defect and genome-wide upd test in case of mosaic paternal upd. Borderline values on a single tissue can be ascertained by the analysis of an additional tissue and techniques, also including SNP array, that may better explore the region. On negative cases, CDKN1C mutations are then searched to complete the BWS molecular testing. Should the clinical diagnosis not be confirmed, differential diagnoses have to be taken into account and whole exome deep sequencing and array-based methods should be considered