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EGA Institute for Women's Health

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Application of Single Molecule, Real-Time sequencing for preimplantation genetic diagnosis and the analysis of embryonic genome activation

Principal Supervisor: Sioban SenGupta

Subsidiary Supervisor: TBC

Project Summary:

Single Molecule Real Time (SMRT) Sequencing is a new technology that enables sequencing from individual DNA molecules without the need for amplification and fragmentation of the target sequence. This sequencing method also allows detection of DNA methylation without bisulfite conversion1,2.

This project will involve the development of SMRT sequencing for direct mutation detection and haplotyping in preimplantation genetic diagnosis to enable genetic analysis in couples with de novo mutations and no previous family history of dominantly transmitted genetic disorders3. The presence of mosaicism in the affected parent can confound diagnostic results therefore the ability to detect mosaicism using SMRT sequencing will be assessed.

Early embryonic development relies on the co-ordinated breakdown of maternal mRNA transcripts that are stored in the oocyte together with the activation of the embryonic genome4,5. In this project SMRT sequencing will also be used to explore the embryonic genome activation of checkpoint and DNA repair genes. There will be a focus on any differential activation of maternally and paternally inherited genomes to determine whether embryo development is differently affected by inherited mutations depending upon the parental origin of the mutation.

References:

  1. Chaisson,M. et al. Resolving the complexity of the human genome using single-molecule sequencing. Nature 2015, 517, 608–611
  2. Flusberg, B. A., et al  Direct detection of DNA methylation during single-molecule, real-time sequencing. Nature Methods, 2010, 7(6), 461–465
  3. SenGupta SB, Delhanty JD. Preimplantation genetic diagnosis: recent triumphs and remaining challenges. Expert Rev Mol Diagn. 2012, 12(6):585-92
  4. Jaroudi S., et al Expression profiling of DNA repair genes in human oocytes and blastocysts using microarrays. Hum Reprod. 2009, 24(10):2649-55
  5. Tulay P, Naja RP, Cascales-Roman O, Doshi A, Serhal P, SenGupta SB.Investigation of microRNA expression and DNA repair gene transcripts in human oocytes and blastocysts. J Assist Reprod Genet. 2015, 32(12):1757-64