1. Application of NGS technologies to Preimplantation Genetic Diagnosis (PGD)
Andreu Paytuví
MSc in Bioinformatics, UAB

2. Little bits of history
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3. Little bits of history
First baby born from NGS!
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4. Why PGD?
Pictures from: Fertility Institute of New Jersey & New York. http://www.center4ivf.com/ (Accessed 28/02/2014).
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5. Why PGD?
Pictures from: Chromosome Screening. http://www.chromosome-screening.org/ (Accessed 28/02/2014).
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Example of aneuploidy:
Turner syndrome

6. Why PGD?
Pictures from: Theisen, A. (2008) Microarray-based comparative genomic hybridization (aCGH). Nature Education 1(1):45
aCGH
2500-3500€ for 6-10 embryos
≈350-400€/embryo
Workflow:
16-24h
10
X Y
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7. PGD based on NGS
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8. PGD based on NGS
Sequencing
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9. PGD based on NGS
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seqtk comp hs37m.fa.gz |
awk '/^[0-9MXY]/{x=$4+$5;y=x+$3+$6;print $1"t"x/y}'
Sequencing
GC bias correction
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10. Data analysis
Picture from: Rico, Alan. Pre-implantation Genetic Testing on Ion Torrent™ PGM™ System. Ion Tour 2013, Life Technologies. Microsoft PowerPoint file.
Based on Hidden Markov Model (HMM):
- States: Ploidy status (monosomy, disomy, etc.).
- Observations: coverage (1X, 2X, 5X, 10X, etc.).
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11. Data analysis
Picture from: Rico, Alan. Pre-implantation Genetic Testing on Ion Torrent™ PGM™ System. Ion Tour 2013, Life Technologies. Microsoft PowerPoint file.
Based on Hidden Markov Model (HMM):
- States: Ploidy status (monosomy, disomy, etc.).
- Observations: coverage (1X, 2X, 5X, 10X, etc.).
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12. Data analysis
Ion PGM™
150K reads
aCGH
Up to 32 embryos per run
≈70€/embryo
Workflow:
≈15h
Pictures from: Rico, Alan. Pre-implantation Genetic Testing on Ion Torrent™ PGM™ System. Ion Tour 2013, Life Technologies. Microsoft PowerPoint file.
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13. Data analysis
CFTR case
Picture from: Treff, N. 2013. Evaluation of targeted next-generation sequencing–based preimplantation genetic diagnosis of monogenic disease. Fertility And Sterility, 99:5, p. 1377-1390.
Picture from: Rico, Alan. Pre-implantation Genetic Testing on Ion Torrent™ PGM™ System. Ion Tour 2013, Life Technologies. Microsoft PowerPoint file.
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14. Prenatal diagnosis
Pictures from: Bianchi, D. W. 2012. From prenatal genomic diagnosis to fetal personalized medicine: progress and challenges. Nat Med., 18, 1041–1051.
Alkan, C., et al. 2010. Personalized Copy-Number and Segmental Duplication Maps using Next-Generation Sequencing. Nat Genet., 41(10), 1061–1067.
Z score of an observation is the number of standard deviations it falls above or below the mean.
-Z score of mean = 0
-Cut-off value of 3 (|Z| > 3)
LOESS method (GC correction)
x --> region
y’ --> corrected counts
f(x) --> mean of counts for that GC%
e(x) --> expected counts (overall average)
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15. Other references:
Recommended talks:
Munne, S., et al. 2005. Preimplantation genetic diagnosis reduces pregnancy loss in women aged 35 years and older with a history of recurrent miscarriages. Fertility and Sterility, 84(2), 331-335.
Handyside, A. H. 2013. 24-chromosome copy number analysis: a comparison of available technologies. Fertility and Sterility, 100(3), 595–602.
Life Technologies, Aneuploidy detection by low-pass whole-genome sequencing on the ion PGM™ system, Appl. Note CO06456 0713.
Chen, E. Z., et al. 2011. Noninvasive Prenatal Diagnosis of Fetal Trisomy 18 and Trisomy 13 by Maternal Plasma DNA Sequencing. PLoS One, 6(7), e21791.
Thanks for your attention!