Topic 5: technologies in genetics Flashcards
What conditions are associated with chromosome abnormalities?
Numerous medical conditions, such as spontaneous abortions, congenital anomalies, birth defects, intellectual disabilities, infertility, cancer, and other pathologies
What does the human karyotype consist of?
46 chromosomes with 22 homologous pairs of autosomes and 1 pair of sex chromosomes (XX for female and XY for male).
What are the two major types of chromosomal abnormalities?
Constitutional abnormality and acquired abnormality.
What are the categories within numerical chromosomal abnormalities?
Monosomy and trisomy.
What are the categories within structural chromosomal abnormalities?
Balanced or unbalanced.
How can constitutional chromosomal abnormalities affect normal development?
Through dosage effect, direct damaging effect, discrepant parental origin, and position effect.
What kinds of chromosomal abnormalities are reported in tumor cells?
Deletions, duplications, and translocations.
What are unbalanced structural rearrangements?
Chromosomal abnormalities with excess or lack of DNA in the genome, affecting dosage-sensitive genes.
What are balanced structural rearrangements?
Not losing or gaining material, just moving it around, including reciprocal translocation, inversion, and Robertsonian translocation.
What is the role of cytogenetic analysis in hematology and oncology?
Identification of abnormalities related to cancer initiation and progression. Important for diagnosis, prognosis, and therapeutic decisions.
What are unbalanced structural rearrangements especially problematic for?
Dosage-sensitive genes, such as haploinsufficient and triplosensitive genes.
What are examples of unbalanced structural rearrangements?
Deletion (terminal or interstitial), duplications (often in tandem), ring and supernumerary marker.
How are chromosomal abnormalities categorized?
Numerical (e.g., monosomy or trisomy) and structural (balanced or unbalanced).
What morphological groups can chromosomes be classified into?
Metacentric, submetacentric, and acrocentric.
What is a dosage effect in terms of chromosomal abnormalities?
A loss or gain of chromosomal material, whether for a whole chromosome or a part of a chromosome, that affects normal development.
What is a direct damaging effect in chromosomal abnormalities?
Disruption of a gene at the breakpoint of a rearrangement, such as a double-stranded break.
What is an effect due to the discrepant parental origin of a chromosome or chromosomal segment?
Known as genomic imprinting, it can lead to conditions such as uniparental disomy, where both copies of a chromosome are inherited from one parent.
What is a position effect in chromosomal abnormalities?
A gene in a new chromosomal environment functions inappropriately, such as when a gene comes close to a strong enhancer.
What is constitutional mosaicism?
A condition where an additional cell line with a different chromosomal complement arises in embryonic or pre-embryonic life and becomes part of the organism.
What is a chimera in terms of chromosomal abnormalities?
An organism derived from two different zygotes, resulting in different cell lines within the same individual.
What is G-banding in chromosome analysis?
G-banding involves trypsin treatment followed by Giemsa staining to produce distinctive and reproducible patterns of transverse light and dark bands along the chromosomes.
What are the benefits and limitations of G-banding?
Benefits: Viewing entire genome, visualizing individual cells and chromosomes, detecting balanced and unbalanced rearrangements.
Limitations: Resolution limit around 5-10 Mb, requires actively growing cells.
What is Fluorescence In Situ Hybridization (FISH)?
A DNA probe labeled with a fluorescent dye hybridizes to the metaphase or interphase chromosome, allowing the study of alterations at the submicroscopic level.
What are the benefits and limitations of FISH?
Benefits: Fast, higher resolution than G-banding, can study non-dividing tissues, shorter TAT for results.
Limitations: Limited to the region of the genome complementary to the probe.
What is Quantitative Fluorescence (QF)-PCR used for?
QF-PCR is used to study multiple DNA polymorphic microsatellite markers on chromosomes 13, 18, 21, X, and Y for rapid detection of common trisomies and monosomy X.
What are the steps in QF-PCR?
DNA extraction, microsatellite marker PCR amplification using fluorescent labeled primer, capillary electrophoresis of fluorescent DNA fragments, analysis of microsatellite loci size and height.
What are the benefits and limitations of QF-PCR?
Benefits: Fast TAT, automated, cost-effective.
Limitations: Targeted approach, will not detect anomalies affecting other chromosomes.
What is Chromosomal Microarray (CMA)?
CMA involves mixing patient DNA and control DNA to hybridize to their complementary sequences on the array, identifying copy number gains and losses across the genome.
What are the benefits and limitations of CMA?
Benefits: Whole genome approach, less reliance on clinician’s suspicion, higher resolution.
Limitations: Cannot identify balanced chromosome rearrangements, cannot establish the location or orientation of chromosome segment duplications.
What factors are considered in the interpretation of CMA data?
It is imperative to distinguish between pathogenic, Variation of Unclear Significance (VUS) and benign CNVs by incorporating family studies, information about the gene content of the region in question, and data from CNV databases and current literature
How is chromosome compaction achieved for chromosome analysis?
By the addition of various proteins (histones, condensins) and arrangement of the chromatin into loops. From interphase to metaphase chromosome, the compaction level goes from 1 to 10,000.
How are metaphase chromosomes obtained for analysis?
Blood is sampled, phytohemagglutinin is added to stimulate lymphocyte division, and mitotic spindle is disabled (e.g., with methotrexate) to stop cell division at mitosis stage.
What are the main steps of the FISH technique?
Denaturation of DNA, hybridization (overnight), post-hybridization washes, and visualization of signals under the fluorescent microscope.
What types of probes are used in FISH?
Centromere-specific and locus-specific probes
What is the role of capillary electrophoresis in QF-PCR? (i.e how do we interpret the data?)
During capillary electrophoresis, amplified fluorescent microsatellite fragments of different lengths migrate at different speeds, and the fluorescence is measured.
How does CMA identify copy number gains and losses?
Red and green intensities are measured, indicating equivalent dosage between the two genomes (yellow) or a relative gain (green) or loss (red) in the patient sample
What are some of the clinical applications of CMA?
First-line test for fetuses with major malformations and patients with unexplained developmental delay, intellectual disability, autism, multiple congenital anomalies, or dysmorphic features.
What are morbid and non-morbid genes in the context of CNVs?
Morbid genes are associated with diseases, while non-morbid genes are not.
what is a microsatellite?
- A microsatellite is a tract of repetitive DNA.
- Microsatellites are present across the genome and the length of the tracts vary between chromosomes and between individuals.
