Next Generation Sequencing

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Next Generation Sequencing

• A paradigm shift practice with great opportunity
  and challenge

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Next Generation Sequencing

• What can NGS do
• Detects nucleotide substitution
• Detects indel
• Detects CNV
• Detects translocation
• Detects inversion
• Detects methylation

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Next Generation Sequencing

• What are involved to do NGS
• Panel design and target capture
• Bar-coding for multiplexing
• Sequencing (choices of platform)
• Informatics infrastructure
• Sequence data analysis pipelines
• Data (Variant interpretation)

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Examples of NGS platforms
100Mb 500 100bp/reads
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Mi-Seq
1-1.5Gb 1200 150bp/run
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Sanger vs. NGS
Sanger sequencing next generation
sequencing
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Analyzing the clinical significance of the
variants
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VAST ARRAY OF GENETIC DISORDERS

• Chromosomal
• Trisomies, aneuploidies, etc.
• Metabolic
• SLO, CDG, many others
• Organ Specific
• Cardiopathies, GI, Renal
• Protein
• DMD, Huntington, Immunoglobulins
• Cancer
• At risk mutations
• And On and On

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Variant categorization

• Known deleterious (condition relevant and
  incidental)
• Presumed deleterious
• VUS (variants of undetermined significance)
• Presumed benign
• Known benign

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Reportability

• Clinically actionable--YES
• Clinically valid but not actionable--YES/NO
• Unknown or no clinical significanceNO
• Presumed benign/known benign--NO

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Counseling

• Pre-test
• Post-test
• Update
• Who gets the test? Who should know the results?
  (part of the results or all)

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448 severe recessive childhood diseases 7717
regions from 437 target genes 93 of target
nucleotide with gt20X coverage 95
sensitivity 100 specificity 104 samples
tested Average carrier burden 2.8 (0-7)