Genetic Testing Flashcards
Chromosomal Analysis
Suspected abnormality of chromosome number or structure (deletion, insertion, rearrangements). Frequently obtained from pregnant women > 35 years (amniocentesis or chorionic villus sampling), from patients with congenital abnormalities (dysmorphisms, structural organ defects, mental and/or growth retardation), from families with multiple miscarriages and/or fertility problems, and directly from certain cancer biopsies.
Chromosomal Analysis can diagnose
aneuploidies (abnormal chromosome number), chromosome deletions, duplications, and insertions of moderate to large size (>3,000-5,000 kb / 3-5 Mb), and rearrangements.
Chromosomal Analysis cannot diagnose
single gene deletions, point mutations, small deletions, duplications, and insertions, methylation defects, trinucleotide repeat abnormalities
FISH
Used to diagnose deletions, some translocations, and abnormalities of copy number. Often used to detect cytogenetic changes that are at or beyond the limits of resolution obtained by high-resolution chromosomal analysis. FISH for duplications works better on cells in interphase than metaphase (metaphase the chromatin is very compact)
FISH can diagnose
recognized microdeletion syndromes, recognized chromosomal rearrangements (in cancers), and gene copy numbers (cancers). Also useful in diagnosing anueploidies (e.g. trisomy 13, 18, 21) in the prenatal setting.
FISH cannot diagnose
deletions, rearrangements that are not specifically tested for (i.e. FISH probes are specifically designed for each condition). FISH is not always able to detect duplications of gene regions. Point mutations and small deletions cannot be diagnosed with this approach.
Examples of Microdeletion Syndromes
Cri-du-chat, Smith-Magenis, DiGeorge (22qdel), Williams syndrome, Wolf-Hirschhorn, Prader-Willi syndrome, Angelman syndrome.
Expression Arrays
Used to test the RNA expression of genes (i.e. which genes are turned ‘on’ or ‘off’). These are semi-quantitative and test the activity of genes (see figure) rather than just the presence or absence of a gene or genetic variant (expression arrays). These have a small, but likely growing role, in oncology
Chromosomal Microarray Analysis (CMA)
These have a big role in clincal genetics currently. These look for chromosomal DNA losses and gains (so called ‘deletion/duplication’ studies). Sometimes this is also called array comparative genomic hybridization (aCGH) analysis.
General Uses and Indications of CMA
CMA has become fairly standard for looking for small genomic deletions/insertions. You can think of this as a superior method to looking for chromosomal gains than losses than traditional chromosomal analysis because the resolution of the CMA is vastly superior to chromosomal analysis. The probe size used these days is between 100-200 Kb so they can pick up smaller changes than can be appreciated by chromosome analysis. Currently, some labs use >~200 Kb for deletions and >~400 Kb for duplications. •
CMA can diagnose
aneuploidies, unbalanced chromosomal rearrangements, chromosome deletions and duplications > 200 Kb and 400 Kb, respectively.
CMA cannot Diagnose
Deletions/Duplications below the resolution of CMA, nucleotide mutations, balanced chromosomal rearrangements
DNA sequencing
Widely used method of mutation detection based on Sanger-Gilbert method. Able to identify the genetic mutation causing many disease conditions
DNA sequencing general uses and indications
Used to identify sequence changes (mutations) in specific genes. In general you need the following:
o You must know or suspect a specific genetic diagnosis
o The gene must have been identified
o The mutation must be detectable by sequencing (deletions, insertions, rearrangements are not always found by sequencing)
o The mutation must be located in a region of the gene that is actually sequenced (promoter and deep-intronic mutations often missed by commercial tests)
DNA sequencing can diagnose
Mutations in known genes (mutation can be previously reported or can be novel), polymorphic variants, small (1 to ~100 nucleotide) deletion/insertions. Ideal for looking at the sequence of a known disease gene •
DNA sequencing cannot diagnose
The technique is very specific, assaying only the region of the gene(s) for which the test has been designed. Frequently, many clinical genetic tests do NOT routinely sequence all parts of a gene (e.g. promoters, introns). This means that although the approach is often very specific, clinical sensitivity is frequently below 100% (this is an important concept to understand). This technique cannot easily detect larger deletions/insertions, rearrangements, and most chromosomal abnormalities.
NextGen DNA Methodology
Uses massively-parallel sequencing of individual DNA molecules and is likely to replace PCR based DNA sequencing within a few years (and is actually already in clinical use as of early 2012).
Genetic testing is used for several purposes:
diagnosis, risk estimation, pregnancy planning, population screening
A genetic test which gives a ‘normal’ (or ‘wild-type’) genetic sequence is often referred to a ‘_______’ by clinicians
negative result
A genetic test which gives an ‘abnormal’ (or ‘mutant’ genetic sequence for example) is often referred to as being ‘‘_______” or ‘‘_______’ by clinicians (again, be wary of patient confusion)
positive
mutant
polymorphism is defined as
a genetic mutation which is present in 1% of the population studied.
informative genetic test result is
one where the information from a genetic test definitively diagnoses or excludes the disease in question. Put another way, an informative genetic test result is one that is reliably either a true positive or true negative result (ruling-in or ruling-out disease/risk, respectively)
non-informative genetic test result usually refers to
a situation where the genetic test result is normal, but it is not possible to definitively exclude disease/disease risk. A non-informative genetic test result leaves open the possibility that an underlying pathogenic mutation exists, but was missed/not detected by the test.
Allelic Heterogeneity refers to the fact that
multiple mutations in a particular gene (or at a particular loci) can cause disease.
Example of allelic heterogeneity
Cystic fibrosis is an autosomal recessive disease caused my mutations in one gene, CFTR. Over 1,000 different mutations have been reported. Cystic fibrosis shows allelic heterogeneity but is genetically homogenous (e.g. NO Genetic Heterogeneity).
Genetic Heterogeneity:
multiple genes (when mutated) associated with the same phenotype
Example of genetic heterogeneity
Hypertrophic cardiomyopathy (HCM) is an autosomal dominant disease caused by mutations in at least 10 different genes. HCM shows both allelic and genetic heterogeneity.
