Genomics in Modern Medicine Flashcards
What is the gold standard of genetic testing?
Sanger PCR
What does sanger PCR do?
Gene by gene, exon by exon sequencing
What are the downfalls of sanger PCR?
- Time consuming
- Limited to known genes/regions
What is next generation sequencing (NGS)?
Technologies that permit rapid interrogation of DNA, up to and including entire genomes, via massively parallel sequencing
What are the benefits of next generation sequencing (NGS)?
Decreasing cost and comparatively rapid results
How many base pairs in the human genome?
3.2 billion base pairs
How many nucleotides in the human genome?
6.4 billion
How many genes in the human genome?
19,000 genes (1-2% of the genome)
How many proteins does the human genome encode?
100,000
What is the normal human variation?
0.5% or 16 million base pairs
What can genetic variation be?
Size
Location
What can genetic variation impact?
- Impact on a codon
- Impact on the protein
- Impact on expression
What is the least frequent genomic variant?
structural change (ex: whole chromosomes)
What is the most frequent genomic variant?
SNV
What are the ways to test/view the human genome for variants?
karyotype
aCGH
SNP array
gemonic sequencing
What are the order of most to least frequent: in-del, CNV, SNV?
SNV
In-del
CNV
Single nucleotide variants account for about ___ percent of all DNA changes?
75%
Single nucleotide variants are found in 1 in every ________ nucleotides
100-300
Insertions and deletions include up to _____ nucleotides
50-100
In-del occur _____ as often as SNV
1/10
____% of In-del are 1-10 nucleotides in length (very short)
90%
*harder to detect b/c so small
What DNA change is the least frequent but large, so affect the most nucleotides?
CNVs
What step must you do if you want to sequence a panal instead of the whole genome?
enrichment
What step is required for all genome sequencing?
library
What are the steps for whole genome sequencing?
- RNA/DNA isolation
- library
- genome
- sequencing
What are the steps for a small section genome sequencing?
- RNA/DNA isolation
- library
- panel
- enrichment
- sequencing
What is the goal of library preparation?
add primers and barcodes to identify individuals
What is shotgun sequencing?
fragmenting DNA during library preparation
Each DNA fragment has what when library is completed?
- Sequencing primer
- Adapters
- Barcode
What is enrichment?
Involves selecting out specific regions of interest for sequencing using capture probes
Why would you sequence a small chunk instead of the whole genome?
- Pooling allows for samples to be mixed together before enrichment
- Allows for more sequencing of targeted regions
- Decreases costs and time per sample
How does sequencing works?
- Fluorescent nucleotides are incorporated one at a time
- Lasers excite the fluorescence, which is read by the instrument
What is variant detection?
the analysis of sequence data to find the differences between an individual’s DNA and the human reference genome
What are the steps in the variant detection process?
- Sequence alignment
- Candidate variant identification
- Consensus calling
- Characterization
What is the variant count for a typical small panel?
8000
What is the variant count for a typical exome?
300,000
What is the variant count for a typical whole genome?
4,000,000
What is the sensitivity and specificity for SNPs?
> 99%
What does next generation sequencing (NGS) allow for?
diagnosis of atypical presentations that would not have otherwise be considered
What are examples of inherited syndromes that affect tooth development?
- Mineralization Defects (PHEX, DMP1, FGF23)
- Ectodermal dysplasias
- Hypo/Oligodontia (PAX9)
- Enamel defects such as Amelogenesis Imperfecta (ENAM)
What is amelogenesis imperfecta?
Several forms of autosomal dominant enamel dysplasia
- Enamel hypoplasia or hypocalcification
How many genes associated with non-syndromic amelogenesis imperfecta?
at least 18
What is the prevalence range of ameologenesis imperfecta?
1/700 to 1/14,000
What were the 3 genes that had variants related to amelogenesis imperfecta?
- Amelogenin
- Amelotin
- Enamelin
What are common barriers to genetic testing in dentistry?
- Identified a need for more education
- Justification of testing
- Changing landscape (Patients will ask for it)
___ permits a new way of molecular diagnosis via panels, exomes, and genomes
NGS
