Term
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Definition
| Quinacrine banding: use fluorescent microscope to band chromosomes; replaced by G-banding |
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Term
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Definition
Giemsa banding: partially digest chromosome proteins with trypsin before staining
400-800 bands/chromosome |
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Term
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Definition
| Reverse banding: use heat treatment; reverses the black and white color; particularly good for the distal end of chromosomes |
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Term
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Definition
| Visualize the constitutive heterochromatin near centromeres |
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Term
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Definition
| Nucleolar Organizing Region stains: stains the satellite and stalk of acrocentric chromosomes |
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Term
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Definition
| Stain the chromosome in prophase or pre-metaphase to identify more, particularly less obvious abnormalities |
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Term
| Fluorescent In Situ Hybridization (FISH) |
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Definition
Use a DNA probe marked with fluorescent dye to hybridize with and identify chromosomes; better than high resolution banding and can identify a 1 million base pair deletion
Detects: extra chromosomes, deleted chromosomes, chromosomal rearrangement |
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Term
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Definition
Uses combinations of 5 dyes to color all the chromosome
Useful identifying small chromosome rearrangements |
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Term
| Comparative Genomic Hybridization (CGH) |
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Definition
1) Extract DNA from test source
2) Label DNA with a color (red)
3) Label DNA from control with a different color (green)
4) Allow both to hybridize with normal metaphase chromosomes
Red = chromosome duplication
Green = chromosome deletion
Limitation: can't detect deletions or duplications less than 5-10 million bp |
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Term
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Definition
Same as CGH except test and control DNA is hybridized with a microarray and probes for specific areas of a chromosome
Detects 1 million bp things
Can't see things less than 150,000 bp unless a smaller probe is used
*Can't detect balanced rearrangements |
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Term
| PCR- Polymerase Chain Reaction |
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Definition
| Amplification of DNA so you can later perform many cytogenetic tests on the amplified DNA |
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Term
| Direct Detection of DNA Alterations |
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Definition
Sequence the DNA, then compare it to a known wild-type DNA sequence to detect mutations
*Use when the alteration for the given disease is known |
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Term
| Microarray-based DNA sequencing |
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Definition
Oligonucleotides for wild-type and mutation sequence are "tiled," labeled with fluorescent dye, and the array is tested for hybridization.
Strong signal = that sequence, and computerized algorithms decode the DNA sequence based on the fluorescent hybridization pattern. |
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Term
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Definition
Indirect approach
These enzymes recognize and cut DNA at specific sequences. If a mutation occurs at a restriction site, add the restriction enzyme, amplify DNA with PCR, and you get two different sized fragments- the mutant one is weird and the normal one is normal sized.
If you run it on agarose gel electrophoresis, the bands travel different distances.
*Use when mutation occurs at an invariable place. |
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Term
| Fluorescent Labels in PCR mixture |
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Definition
Indirect Method
Add fluorescently labeled nucleotides to the mix. The normal sequence will have the correctly incorporated base, whereas the mutant will incorporate a different base.
*Can detect mutant DNA in heterogenous mixtures of normal and abnormal cells
*Use on prenatal samples, peripheral blood lymphocytes, cancer biopsies, tissue sections
*Use to detect chromosomal abnormalities, microdeletions, translocations, gene amplification, mapping newly isolated genes of interest |
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Term
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Definition
Indirect Method
Use fluorophores in "real time" so you don't have to digest fragments and do elecctrophoresis- save time! |
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Term
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Definition
| Add primers that go on either side of a known segment that would have variable size, like Fragile X. The PCR products will be variable sizes, and you can determine the problem- size difference determined by running products on gel |
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Term
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Definition
Assesing a marker loci in family members having the disease or trait of interest, assuming the marker loci is close to the disease allele transmitted through pedigrees
*Use if you don't know what the gene is, prenatal and presymptomatic diagnosis, or several different mutations cause the diseased state |
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Term
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Definition
Genetic marker that can be followed from parent to child
- prevalent throughout genome and stable
*Used in linkage analysis for identifying haplotypes associated with disease |
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Term
| Repeat-length polymorphisms |
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Definition
Short repetitive sequences of DNA
Microsatellite repeats: <1000 bp, 2-6 bp repeat size
*Ideal for differentiating two individuals and following transmission of markers from parent to child
Minisatellite repeats: 1000-3000 bp repeats, repeat motif of 15-70 bp |
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Term
| Genome-Wide Association Studies (GWAS) |
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Definition
| Large number of patients have their genomes examined for variants, which allows the identification of variable areas in the genome or areas where a disease is likely to occur- the candidate gene |
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Term
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Definition
Hybridize radiolabeled sequence-specific probes to genomic DNA that was digested with a restriction enzyme and separated by gel electrophoresis- need a normal control
*Use to detect changes in the structure of specific loci
*Use for large trinucleotide expansion diseases (Fragile X) and clonal Ig gene rearrangements (lymphoma) |
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Term
| Array Comparative Genomic Hybridization (CGH) |
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Definition
Label normal and test samples with dye (CY5 and CY3), hybridize samples with DNA probe- equal expression = yellow, overproduction of mutation is red (CY3) meaning duplication, underproduction is green (CY5) meaning deletion
*Use to detect abnormalities without prior knowledge of the problem
*Localizes amplifications and deletions in the test sample |
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Term
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Definition
| Study of heritable chemical modifications of DNA or chromatin without altering the DNA sequence- DNA methylation, histone methylation and acetylation |
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Term
| Sodium Bisulfite Treatment |
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Definition
| Treat DNA with this, converts unmethylated cytosines to uracil, methylation-specific PCR is then performed |
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