Preparing a karyotype Harvest cells Postnatal diagnostic karyotype tumor biopsy skin cells from mouth (ie for non-cancer related diagnoses) Prenatal diagnostic karyotype chorionic villi sampling (CVS) amniocentesis
Who is offered amniocentesis or CVS? Maternal age (women 35 or older) Risk of Downs syndrome: mother in 20s 1/ % OK mother at 35 1/ % OK mother at 40 1/100 99% OK
Downs syndrome: how does it happen? Chromosomal non-disjunction during meiosis of eggs and sperm. 1. Chromosomes replicate 2. Homologous chromosomes separate 3. Chromatids from each chromosome separate
Who is offered amniocentesis or CVS? Maternal age (women 35 or older) Risk of Downs syndrome: mother in 20s 1/ % OK mother at 35 1/ % OK mother at 40 1/10099% OK A previous child or pregnancy with a birth defect Screening test with a positive result Other family history
Prenatal diagnosis: amniocentesis Sampling cells from amniotic fluid Usually done ~ 15–18 weeks
Prenatal diagnosis: chorionic villi sampling (CVS) Sampling cells from placenta Usually done 10–12 weeks
Preparing a karyotype Harvest cells Culture cells 1–2 days Arrest cells in metaphase with colchicine metaphase
Mitosis DNA replication chromosomes condense nuclear envelope breaks down chromosomes aligned on spindle fibres
Preparing a karyotype Harvest cells Culture cells 1–2 days Arrest cells in metaphase with colchicine Spread cells on slide and stain Count chromosomes in 20 representative cells Capture image of five best cells and construct karyotypes for each metaphase
Metaphase spread chromosomes stained with DAPI, a fluorescing stain that specifically binds double-stranded DNA FISH analysis of chromosomes: Fluorescent In Situ Hybridization
FISH Expose DAPI-stained metaphase chromosomes to fluorescent probes red = control probe for centromere of the X chromosome and another probe for end of chromosome X green = probe for the end of chromosome 4
DiGeorge syndrome/CATCH22 Microdeletion on chromosome 22 Birth defect that affects the immune system Absence or underdevelopment of the thymus and parathyroid glands Facial features include low-set ears, wide-set eyes, small jaw and bowing up of upper lip
FISH tests: DiGeorge syndrome Expose DAPI-stained chromosomes to mixture of fluorescent probes green = control probe for chromosome 22 red = probe for DiGeorge region on long arm of chromosome 22
FISH tests: Painting chromosomes Expose chromosomes to fluorescent probes that highlight entire chromosomes.
FISH tests: Painting chromosomes green = chromosome 13 red = chromosome 21 aqua = chromosome 18 green = X chromosome red = Y chromosome nuclei from the same foetus Expose chromosomes to fluorescent probes that highlight chromosomes 13, 18, 21, X and Y.
Trait A physical characteristic that is determined by genes, eg eye colour.
Thumb shape Earlobe attachment Human traits hhHh or HH Hitchhikers thumb AA or Aaaa unattached attached
Thumb shape Human traits hhHh or HH Hitchhikers thumb Genotype vs phenotype Genotype = specific allelic make-up of an individual, eg HH, Hh or hh Phenotype = an observable physical or measurable biochemical characteristic, eg thumb shape or lack of a particular enzyme
Punnett squares Remember these?? Used to determine the probability of an offspring having a particular genotype H allele = dominant h allele = recessive H HhHh hhHh or HH Hitchhikers thumb
Punnett squares Now try it backwards H allele = dominant h allele = recessive hhHh or HH Hitchhikers thumb hh Hh hh
Recombination: Shuffling the deck DNA crossovers in chromosome pairs that result in children receiving a different combination of genes than either parent