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  1. 2. <ul><li>The study of chromosome and the related disease states caused by abnormal chromosome number andor structure. </li></ul><ul><li>Chromosomes : complex structures located in the cell nucleus, composed of DNA, histone and non-histone proteins, RNA, and polysacchairdes. </li></ul><ul><li>Cytogenetics involves the study of chromosomes. </li></ul><ul><li>Both the number and structure of the chromosomes are analyzed. </li></ul>
  2. 3. <ul><li>A tissue that can be stimulated to undergo cell division in-vitro </li></ul><ul><li>It is only during mitosis of the cell cycle that distinct chromosomes can be visualized with a light microscope </li></ul><ul><li>Examples of tissues that can be studied: chorionic villi, amniotic fluid, peripheral blood (lymphocytes), skin (fibroblasts), bone marrow </li></ul>
  3. 4. <ul><li>Cells are stimulated to divide and are incubated for 48-96 hrs. </li></ul><ul><li>Cells are arrested at mitosis </li></ul><ul><li>Cells are “dropped” onto glass slides and G-banded </li></ul>
  4. 6. <ul><li>Each chromatid represents one molecule of DNA </li></ul>
  5. 7. Effect Area Stained Stain Type Under UV light, distinct fluorescent banded pattern for each chromosome. Chromosome arms; mostly repetitive AT-rich DNA Quinacrine Q-banding Distinct banded pattern for each chromosome; same as Q-banding pattern except single additional band near centromere of chromosomes 1 and 16 Chromosome arms; mostly repetitive AT-rich DNA Giemsa G-banding Reverse banding pattern of that observed with Q- or G-banding Chromosome arms; mostly unique GC-rich DNA Variety of techniques R-banding Largest bands usually on chromosomes 1, 9, 16, and Y; chromosomes 7, 10, and 15 have medium-sized bands; size of C-bands highly variable from person to person Centromere region of each chromosome and distal portion of Y chromosome; highly repetitive, mostly AT-rich DNA Variety of techniques C-banding
  6. 8. Non-Banded Karyotype
  7. 10. <ul><li>Chromosomes can be identified by: </li></ul><ul><li>Their size </li></ul><ul><li>Their shape (the position of the centromere) NB Chromosomes are flexible </li></ul><ul><li>Banding patterns produced by specific stains (Giemsa) </li></ul><ul><li>Chromosomes are analysed by organising them into a KARYOTYPE </li></ul>©
  8. 12. <ul><li>Short arm of the chromosome = p </li></ul><ul><li>Long arm of the chromosome = q </li></ul><ul><li>Bands are numbered independently on the short and long arms </li></ul>
  9. 14. 7 18 Low/High Resolutions Karyotype
  10. 16. Normal Karyotype/ Ideogram
  11. 17. Q-Banding
  12. 18. C-Banding
  13. 19. R - Banding
  14. 20. <ul><li>Advantages </li></ul><ul><li>1- Enable the entire genome to be viewed at one time. </li></ul><ul><li>2- Suitable when a specific anomaly is suspected ( e.g. Philadelphia in CML ) and as a general diagnostic tool to detect additional chr. Abnormalities commonly seen in disease progression of CML. </li></ul><ul><li>Disadvantages </li></ul><ul><li>1- Detect major structural abnormalities </li></ul><ul><li>( one band = 6mb of DNA ~ 150 genes ). </li></ul><ul><li>2- Labor intensive and highly dependent upon operator experience and skills. </li></ul>Advantages and Disadvantages of conventional technique
  15. 21. <ul><li>46,XY </li></ul><ul><li>46,XX </li></ul><ul><li>47,XX,+21 </li></ul><ul><li>45,XO </li></ul><ul><li>46,XY,5p- </li></ul><ul><li>46,XX,t(9;22) </li></ul><ul><li>46,XY,del(15q11-q13) </li></ul>
  16. 22. <ul><li>Chromosomes can be identified by: </li></ul><ul><li>Their size </li></ul><ul><li>Their shape (the position of the centromere) NB Chromosomes are flexible </li></ul><ul><li>Banding patterns produced by specific stains (Giemsa) </li></ul><ul><li>Chromosomes are analysed by organising them into a KARYOTYPE </li></ul>©
  17. 23. <ul><li>A) Polypoidy – change in complete sets of chromosomes (3n, 4n, etc) </li></ul><ul><li>plants > animals. </li></ul><ul><li>Autopolyploidy vs Allopolyploidy </li></ul><ul><li>Plants  more yeild </li></ul><ul><li>Animals  parthenogenesis </li></ul><ul><li>B) Aneuploidy – change in the no. of chromosomes </li></ul><ul><li>nullisomy 2n-2 </li></ul><ul><li>monosomy 2n-1 </li></ul><ul><li>trisomy 2n+1 </li></ul><ul><li>tetrasomy 2n+2 </li></ul><ul><li>Gene dosage effect </li></ul><ul><li>1 - Sex-chromosomal aneuploids . </li></ul><ul><li>2 - Autosomal aneuploids . </li></ul>
  18. 26. <ul><li>Chromosome rearrangements: </li></ul><ul><li>Effects </li></ul><ul><li>Deletion. pseudodominance </li></ul><ul><li>Duplication. gene dosage </li></ul><ul><li>Inversion – paracentric positional </li></ul><ul><li>pericentric </li></ul><ul><li>Translocation. positional </li></ul><ul><li>new linkage rearrangement </li></ul>
  19. 27. <ul><li>Extra single chromosomes or missing single chromosomes </li></ul><ul><ul><li>2N + 1 </li></ul></ul><ul><ul><li>2N - 1 </li></ul></ul><ul><li>Suffix: “-somy” or “-somic” </li></ul><ul><ul><li>2N + 1 = trisomy </li></ul></ul><ul><ul><li>2N - 1 = monosomy </li></ul></ul><ul><ul><li>2N + 2 = tetrasomy </li></ul></ul>
  20. 28. <ul><li>Generally arise through non-disjunction at meiosis </li></ul><ul><ul><li>homologues or chromatids do not separate </li></ul></ul><ul><ul><li>gametes contain 2 or no copies of one chromosome </li></ul></ul>
  21. 30. <ul><li>Most aneuploidies in humans lead to such drastic effects, the fetus is spontaneously aborted early in development </li></ul><ul><li>A few survive ‘til birth; some beyond </li></ul>
  22. 31. <ul><li>47, +21 </li></ul><ul><ul><li>1/700 live births </li></ul></ul><ul><ul><ul><li>over 60% of conceptions aborted spontaneously </li></ul></ul></ul><ul><ul><ul><li>20% stillborn </li></ul></ul></ul><ul><ul><ul><li>incidence increases sharply w/ maternal age </li></ul></ul></ul><ul><ul><ul><ul><li>1/300 for 35 year olds </li></ul></ul></ul></ul><ul><ul><ul><ul><li>1/22 for 45 year olds </li></ul></ul></ul></ul>
  23. 32. <ul><ul><li>characteristic facial appearance </li></ul></ul><ul><ul><ul><li>small nose, flat face, epicanthal fold </li></ul></ul></ul><ul><ul><li>single palmar crease </li></ul></ul><ul><ul><li>mental retardation (avg. IQ < 50) </li></ul></ul><ul><ul><li>multiple complications </li></ul></ul><ul><ul><ul><li>heart disease </li></ul></ul></ul><ul><ul><ul><li>leukemia </li></ul></ul></ul><ul><ul><ul><li>epilepsy </li></ul></ul></ul><ul><ul><li>some fertility </li></ul></ul>
  24. 33. <ul><li>47; +18 </li></ul><ul><ul><li>1/8000 live births; maternal age affect </li></ul></ul><ul><ul><li>low birth weight </li></ul></ul><ul><ul><li>multiple dysmorphic features </li></ul></ul><ul><ul><ul><li>chin, ears, single palmar crease, clenched hands </li></ul></ul></ul><ul><ul><li>malformations of the brain, heart, kidneys, and other organs </li></ul></ul><ul><ul><li>rarely survive beyond 1 year </li></ul></ul>
  25. 34. <ul><li>47; + 13 </li></ul><ul><ul><li>1/20,000 live births; maternal age effect </li></ul></ul><ul><ul><li>multiple dysmorphic features </li></ul></ul><ul><ul><ul><li>micropthalmia, cleft palate, clenched fists, polydactyly, ears and scalp abnormal… </li></ul></ul></ul><ul><ul><ul><li>heart defects; systemic defects…. </li></ul></ul></ul><ul><ul><li>50% die in first month; rarely survive beyond 1 year </li></ul></ul>
  26. 35. <ul><li>1/500 live male births (?) </li></ul><ul><li>often asymptomatic except for sterility, learning disabilities </li></ul><ul><li>small testes; low testosterone levels </li></ul><ul><li>poorly developed male 2 o sexual charact. </li></ul><ul><ul><li>some female characteristics: </li></ul></ul><ul><ul><ul><ul><li>enlarged breasts, elongated limbs, increased incidence of “female” diseases: breast cancer, scoliosis, osteoporosis </li></ul></ul></ul></ul><ul><ul><li>hormone therapy improves symptoms </li></ul></ul>
  27. 36. <ul><li>1/1000 live male births? </li></ul><ul><li>formerly called “criminal chromosome” </li></ul><ul><ul><li>99% asymptomatic, though high incidence in penal institutions for the mentally subnormal (20/1000) </li></ul></ul><ul><ul><li>lower than average intelligence? (learning disabilities) above average height, tendency to severe acne </li></ul></ul>
  28. 37. <ul><li>1/2500 live female births </li></ul><ul><li>generally asymptomatic ‘til puberty </li></ul><ul><ul><li>lack of 2 o sexual characteristics; amenorrhoea </li></ul></ul><ul><ul><li>short stature; low hair line; </li></ul></ul><ul><ul><li>heart disease, renal malformations, ovaries generally underdeveloped, sterile </li></ul></ul><ul><ul><li>hormone therapy helpful </li></ul></ul>
  29. 38. <ul><li>1/700 live female births (?) </li></ul><ul><li>generally asymptomatic </li></ul><ul><ul><li>15 - 25% mildly mentally retarded </li></ul></ul><ul><ul><li>some sterility </li></ul></ul>
  30. 39. <ul><li>Changes in the numbers of genes </li></ul><ul><ul><li>deletions </li></ul></ul><ul><ul><li>duplications </li></ul></ul><ul><li>Changes in the location of genes </li></ul><ul><ul><li>inversions </li></ul></ul><ul><ul><li>translocations </li></ul></ul><ul><ul><li>Robertsonian changes </li></ul></ul>
  31. 40. <ul><li>Loss of a (generally small) segment of chromosome </li></ul>A B C D E F G A B D E F G C
  32. 41. <ul><li>Arise through spontaneous breakage </li></ul><ul><ul><li>some chromosomes have fragile spots </li></ul></ul><ul><ul><li>radiation, UV, chemicals, viruses may increase breakage </li></ul></ul>
  33. 42. <ul><li>May arise through unequal crossing over </li></ul>A B C D E F G A B C D E F G x A B C D E F F G A B C D E G Deletion Duplication
  34. 43. <ul><li>Large deletions will most probably be lethal </li></ul><ul><li>Smaller deletions may allow survival </li></ul><ul><ul><li>E. coli : deletions of up to 1% have been observed in living cells </li></ul></ul><ul><ul><li>D. melanogaster : deletions of up to 0.1% observed </li></ul></ul>
  35. 44. <ul><li>Cri-du-chat syndrome </li></ul><ul><ul><li>Micro deletion of chromosome 5 </li></ul></ul><ul><li>DiGeorge syndrome </li></ul><ul><ul><li>Micro deletion of chromosome 22 </li></ul></ul><ul><li>Angelman syndrome </li></ul><ul><ul><li>Micro deletion of chromosome 15 </li></ul></ul><ul><li>Prader-Willi syndrome </li></ul><ul><ul><li>Micro deletion of chromosome 15 </li></ul></ul>
  36. 45. <ul><li>lack of muscle tone in newborn </li></ul><ul><li>poor swallowing reflex </li></ul><ul><li>as adult - gross obesity </li></ul><ul><li>mean I.Q. ~ 50 </li></ul><ul><li>microdeletion of 15 </li></ul><ul><li>developmentally delayed </li></ul><ul><li>jerky movements </li></ul><ul><li>stiff, fixed smile </li></ul><ul><li>uncontrolled laughter </li></ul><ul><li>abnormal E.E.G., epilepsy </li></ul><ul><li>microdeletion of 15 </li></ul>Prader-Willi Angelman
  37. 46. <ul><li>Redundant segment of a chromosome </li></ul>A B C D E F G A B C D E F F G
  38. 48. <ul><li>180 o reversal of chromosome segment </li></ul>A B C D E F G H I J K A B C H G F E D I J K 180 O
  39. 49. <ul><li>Produced through breakage and reassociation of chromosome </li></ul>A B C D E F G
  40. 50. <ul><li>Produced through breakage and reassociation of chromosome </li></ul>A B C D E F G
  41. 53. <ul><li>May change phenotype through “position effects” </li></ul><ul><ul><li>move active genes to sites generally inactive; lose gene function </li></ul></ul><ul><ul><li>move inactive genes to sites generally active; gain gene function </li></ul></ul><ul><li>May act to preserve blocks of genes (specific alleles) which function well together </li></ul>
  42. 54. <ul><li>Exchange of segments between non-homologous chromosomes </li></ul>F A B C D E L M N O P Q
  43. 55. A B C D E F O N M L Q P Remember: There are still two normal chromosomes in the cell! = translocation heterozygote
  44. 56. Translocation
  45. 57. <ul><li>Balanced </li></ul><ul><li>Unbalanced </li></ul><ul><li>Lead to impaired fertility </li></ul><ul><ul><li>complications to synapsis and segregation </li></ul></ul><ul><li>May lead to changes in phenotype </li></ul><ul><ul><li>position effects </li></ul></ul><ul><ul><li>human cancers and translocations </li></ul></ul>
  46. 58. <ul><li>Several human cancers are associated with reciprocal translocations </li></ul><ul><ul><li>Chronic myelocytic leukemia, many acute leukemias </li></ul></ul><ul><ul><ul><li>Philadelphia chromosome </li></ul></ul></ul><ul><ul><li>Burkitt’s lymphoma </li></ul></ul><ul><ul><ul><li>three translocations </li></ul></ul></ul>
  47. 59. <ul><li>All these involve oncogenes </li></ul><ul><ul><li>oncogenes: genes involved in cell proliferation </li></ul></ul><ul><ul><ul><li>normally active only at specific times </li></ul></ul></ul><ul><ul><ul><ul><li>embryonic/fetal development, etc. </li></ul></ul></ul></ul><ul><ul><ul><li>shut off under normal conditions </li></ul></ul></ul>
  48. 60. Reciprocal translocation between chr. 22 and chr. 9 9 9 22 22
  49. 61. Reciprocal translocation between chr. 22 and chr. 9 9 9 22 22 Chr. 9 segment has an oncogene; when moved, it becomes active
  50. 62. <ul><li>Translocations between chr. 8 and one of three others: </li></ul><ul><ul><li>8 & 2 or 8 & 14 or 8 & 22 </li></ul></ul><ul><li>Chr. 8 has an oncogene </li></ul><ul><li>Chr. 2, 14, 22 have genes coding for antibody production and their enhancer genes </li></ul><ul><li>Oncogene becomes highly active under control of enhancers! </li></ul>
  51. 63. <ul><li>Fusions </li></ul><ul><ul><li>two chromosomes join to form one </li></ul></ul><ul><li>Fissions </li></ul><ul><li>one chromosome splits to form two </li></ul>
  52. 64. Roberstsonian translocations
  53. 65. <ul><li>Familial Down Syndrome </li></ul><ul><ul><li>fusion of chromosomes 14 and 21 </li></ul></ul>14 21 14+21 Carrier; phenotypically normal produces normal, carrier, and Down syndrome children
  54. 66. synapsis Three possible disjunctions
  55. 68. X X
  56. 69. X X X X
  57. 70. © 2007 Paul Billiet ODWS Images believed to be in the Public Domain 23 unpaired chromosomes 23 unpaired chromosomes 23 unpaired chromosomes 23 unpaired chromosomes Fertilisation Child Father 23 pairs of chromosomes Sex cells Meiosis Mother 23 pairs of chromosomes 23 pairs of chromosomes
  58. 71. <ul><li>A special type of cell division </li></ul><ul><li>Used to make sex cells </li></ul><ul><li>Meiosis halves the numbers of chromosomes </li></ul><ul><li>Meiosis picks one chromosome from each pair at random and places them in a sex cell. This results in enormous variation amongst the sex cells. </li></ul>© 2007 Paul Billiet ODWS
  59. 72. Y X X X Fertilisation Possible children Father Sex cells Meiosis Mother XX XY Chance of a girl 50% Chance of a boy 50% © 2007 Paul Billiet ODWS X Y X XX XY X XX XY
  60. 73. <ul><li>The sex of many animals is determined by genes but on chromosomes called sex chromosomes </li></ul><ul><li>The other chromosomes are called autosomes </li></ul><ul><li>One sex is homogametic </li></ul><ul><li>The other sex is heterogametic </li></ul>© 2007 Paul Billiet ODWS
  61. 74. © 2007 Paul Billiet ODWS HOMOGAMETIC SEX HETEROGAMETIC SEX SEX DETERMINATION Female XX Male XY Presence of Y-chromosome = maleness (mammals and fish) Presence of second X-chromosome = femaleness (Drosophila, the fruit fly) Male ZZ Female ZW Birds, amphibians, reptiles, butterflies, moths. Female XX Male Xo Grasshoppers
  62. 75. <ul><li>Constant for each cell in the body (except sex cells which only have half sets). </li></ul><ul><li>Constant throughout the life of an individual (you don’t lose or gain chromosomes) </li></ul><ul><li>Constant for all members of a species </li></ul>© 2007 Paul Billiet ODWS
  63. 76. Mouse Maize © A. Lane Rayburn Image believed to be in the Public Domain
  64. 77. © 2007 Paul Billiet ODWS Organism Chromosome numbers Human 46 Chimpanzee 48 House Mouse 40 Maize 20
  65. 78. Image believed to be in the Public Domain