Array Comparative Genomic Hybridisation and Droplet Digital PCR Uncover Recurrent Copy Number Variation of the TTN Segmental Duplication Region

Genes (Basel). 2022 May 19;13(5):905. doi: 10.3390/genes13050905.


Intragenic segmental duplication regions are potential hotspots for recurrent copy number variation and possible pathogenic aberrations. Two large sarcomeric genes, nebulin and titin, both contain such segmental duplication regions. Using our custom Comparative Genomic Hybridisation array, we have previously shown that a gain or loss of more than one copy of the repeated block of the nebulin triplicate region constitutes a recessive pathogenic mutation. Using targeted array-CGH, similar copy number variants can be detected in the segmental duplication region of titin. Due to the limitations of the array-CGH methodology and the repetitiveness of the region, the exact copy numbers of the blocks could not be determined. Therefore, we developed complementary custom Droplet Digital PCR assays for the titin segmental duplication region to confirm true variation. Our combined methods show that the titin segmental duplication region is subject to recurrent copy number variation. Gains and losses were detected in samples from healthy individuals as well as in samples from patients with different muscle disorders. The copy number variation observed in our cohort is likely benign, but pathogenic copy number variants in the segmental duplication region of titin cannot be excluded. Further investigations are needed, however, this region should no longer be neglected in genetic analyses.

Keywords: Droplet Digital PCR; TTN; copy number variation; segmental duplication; titin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Connectin / genetics
  • DNA Copy Number Variations* / genetics
  • Genomics
  • Humans
  • Muscle Proteins
  • Polymerase Chain Reaction
  • Segmental Duplications, Genomic* / genetics


  • Connectin
  • Muscle Proteins
  • TTN protein, human
  • nebulin

Grants and funding

This research was funded by Magnus Ehrnrooth foundation, Finska Läkaresällskapet, Svenska Kulturfonden, Stiftelsen Dorothea Olivia, Karl Walter och Jarl Walter Perlkéns minne, and Folkhälsan Research Foundation.