Types of genetic tests 1. Cytogenetic 2. DNA 3. Metabolic

Karyotype Picture of the chromosomes in a cell used to check for abnormalities Prenatal diagnosis: trisomy 21 (Down’s syndrome)

Postnatal diagnosis:

Postnatal diagnosis: detecting cancer

Preparing a karyotype Harvest cells (from where?) Postnatal diagnostic karyotype Prenatal diagnostic karyotype

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 Down’s syndrome: mother in 20s 1/1250 99.92% OK mother at 35 1/400 99.75% OK mother at 40 1/100 99% OK

Down’s 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 Down’s syndrome: mother in 20s 1/1250 99.92% OK mother at 35 1/400 99.75% OK mother at 40 1/100 99% 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 metaphase chromosomes condense DNA replication nuclear envelope breaks down metaphase 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

FISH analysis of chromosomes: Fluorescent In Situ Hybridization Metaphase spread chromosomes stained with DAPI, a fluorescing stain that specifically binds double-stranded DNA

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 Expose chromosomes to fluorescent probes that highlight chromosomes 13, 18, 21, X and Y. nuclei from the same foetus green = chromosome 13 red = chromosome 21 aqua = chromosome 18 green = X chromosome red = Y chromosome

Trait A physical characteristic that is determined by genes, eg eye colour.

Human traits Thumb shape hh Hh or HH Earlobe attachment AA or Aa aa ‘Hitchhiker’s thumb’ Earlobe attachment AA or Aa aa unattached attached

Human traits Genotype vs phenotype Thumb shape hh Hh or HH ‘Hitchhiker’s 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 hh Hh or HH H H ‘Hitchhiker’s thumb’ H h H allele = dominant h allele = recessive

Punnett squares Now try it backwards hh Hh or HH Hh Hh ‘Hitchhiker’s thumb’ Hh Hh H allele = dominant h allele = recessive hh hh

Recombination: Shuffling the deck DNA crossovers in chromosome pairs that result in children receiving a different combination of genes than either parent