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22 Cards in this Set
- Front
- Back
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FISH
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Fluorescence in situ hybridization
-labeled probe to deletion region -detect microdeletions -must denature |
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CGH
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Comparative genomic hybridization
-Microarray -different fluorescence for patient and normal DNA -Detect microdeletions/duplications |
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Restriction endonuclease
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Restriction fragment length polymorphism (RFLP)
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Electrophoresis
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Agarose: DNA up to 30kb
Acrylamide: 5-500bp |
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PCR
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-small size amplification
-Fast, use DNA or RNA -Only needs one cell -detect specific mutation alleles w/ mutant primer -CONs: contamination, may not be 100% specific |
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Southern blot
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electrophoresis transfer onto nylon membrane
-hybridize w/ labeled probe -detect large deletions/insertions, point mutations, polymorphisms, methylation, DNA rearrangement -Con: needs lots of DNA and time |
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Allele-specific oligonucleotide
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PCR-amplify patient DNA at common mutant allele fragments
-immobilize oligos w/ mutant alleles on solid -biotin-tagged DNA hybridize to oligos. -detect w/ peroxidase |
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DNA sequencing
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Targeted only. Expensive and time-consuming
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What makes a good screen?
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-window of intervention
-high prevalence -sensitivity (detect all positives), specificity (eliminate negatives) should be high |
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Sensitivity
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A/(A+C)
-test positive divided by all disease |
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Specificity
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B/(B+D)
-test negative divided by all non-disease |
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Conditions for prenatal screening
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-Adv Mat Age
-Previous child w/ anomaly -Carrier / X-linked -Teratogen exposure in mother |
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Alpha fetoprotein
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Maternal serum screen
-TRISOMY 21, DOWN SYNDROME (low) -Hepatically-derived protein in fetal life leaks into maternal blood -False-positive possible due to younger-than-expected fetus -ID neural tube defects (high) |
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Ultrasonography
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Transabdominal-deep penetration, mid-trimester
Transvaginal-low penetration, first trimester -use to DATE the pregnancy -use to detect Teratology (teratology is not chromosomal defect--do not use CVS, amnio) |
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Chorionic Villus Sampling
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Obtain trophoblasts
10-13 weeks gestation |
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Amniocentesis
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Needle through abdominal/uterine walls retrieve amnionic fluid
-15-20 week post-gestation -NEURAL TUBE defects |
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Percutaneous umbilical venous sampling
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Fetal blood sampling in 3rd trimester
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PGD
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Pre-implantation genetic diagnosis
-remove 1-2 cells from 8-cell stage -PCR, FISH |
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Karotype
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Taken in Metaphase
Arranged in order of size. |
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Indications for Chromosomal testing
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-suspected syndrome
-multiple congenital malformations -mental retardation -short stature in girls -leukemia/lymphoma -Adv Mat Age |
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Reverse "Line" blot assay
Linear Array |
Multiplex PCR (multiple primers)
-hybridize w/ oligos WT or mutant -hybrids are colorimetric--Bar Code -ID heterozygote, homozygote, compound heterozygotes |
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Down Syndrome Prenatal diagnostics
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1st Trimester
-free hCG high, plasma protein A low -Ultrasound 2nd Trimester -AFP (reduced), hCG (high), inhibin-A (high) |
