applied lectures Flashcards
Nondisjunction
Meiosis I: failure of two homologous chromosomes to pass to separate cells
Meiosis II: failure of two chromatids of a chromosome to pass to seperate cells
Aneuploidy
abnormal number of certain chromosomes
-when individuals have too many or too few of specific chromosomes
Karyotype and how to make one
organized display of the homologous pairs of chromosomes and the sex chromosomes (46 chromosomes, 22 pairs of autosomes, 2 sex chromosomes (X and Y), each of the chromosomes consists of 2 sister chromatids lying close together)
-useful in genetic screening to identify specific chromosomal defects in their number, size and type
-stop at metaphase
Make one: draw blood (10/20ml), add blood and phytohemagglutinin to stimulate mitosis, incubate at 37C for 2/3 days, add colcemid to culture for 1 to 2 hours to stop mitosis in metaphase, transfer cells to tube, transfer to tube containing fixative, centrifuge to concentrate cells - add low salt solution to eliminate red blood cells and swell lymphocytes, drop cells onto microscope slide and stain w/giemsa, examine with microscope, digitized chromosome images processed to make karyotype
Amniocentesis for testing genetic disorders in fetus
1) amniotic fluid sample is taken at 14-16 week of pregnancy
2) biochemical tests can be performed immediately on the fluid or later cultured tests
3) fetal cells must be cultured for several weeks to obtain sufficient numbers for karyotyping (testing for chromosomal disorders)
-more expensive, takes longer
Chorionic villus sampling (CVS)
1) a sample of chorionic villus tissue can be taken as early as the 8th to 10th week of pregnancy
2) karyotyping and biochemical tests can be performed on the fetal cells immediately, providing results within a day or two
-quick, cheaper
how to make a karyotype w/hypotonic solution
1) blood culture is centrifuged to separate the blood cells from the culture fluid
2) the fluid is discarded and a hypotonic solution is mixed with the cells. This makes the red blood cells swell and burst. The white blood cells swell but do not burst, and their chromosomes spread out
3) another centrifugation step separates the swollen white blood cells. The fluid containing the remnants of the red blood cells is poured off. A fixative (preservative) is mixed with the white blood cells. A drop of the cell suspension is spread on a microscope slide, dried, and stained.
4) the slide is viewed with a microscope, and images of the metaphase chromosomes are sorted by size and shape on a computer
5) the resulting display is the karyotype
Klinefelter Syndrome
47, XXY
-male have an extra X number of sex chromosomes, XXY
-very small testes, and are sterile
-feminine body contours (enlargement of breast tissue)
-normal intelligence
Trisomy
-individuals have 3 copies of a certain chromosome (type of aneuploidy)
ex: down syndrome (three copies of chromosome 21)
-shorter in structure
-pronounced hand ridge
Patau Syndrome (trisomy)
47, 13+
-trisomy of chromosome 13
-severely mentally challenged
-cleft palate (not everyone with cleft lip has patay
-deaf
-malformed organs
Edwards Syndrome
-trisomy of chromosome 18
-small newborns
-low set ears, webbed neck, receding chin
-organ malformations
Turner Syndrome
45 0X,X0,XO (It is characterized by the partial or complete absence of one of the X chromosomes.)
-alteration of sex chromosomes
-a viable human monosomy
-short stature
-webbed neck shield like chest
-internal sex organs do not mature
-females are sterile
in females
Triple X syndrome
XXX
-usually normal phenotype and fertile, other have variable expression
-more introverted/shy
Jacobs Syndrome
alterations of sex chromosomes
XYY
-normal male but much taller than average
-normal fertility and intelligence
why do meiotic errors occur?
they are accidental
-occur w/o any genetic predisposition
-larger percentage of errors occur due to maternal errors (frequency increases with age)
relationship between aneuploidy and cancer
-mitotic chromosome instability is a characteristic of many cancer cells
-relationship of aneuploidy and tumor formation may lead to activation of oncogenes and inactivation of tumor suppressors
Ghulam Mufti
test. cancer cells by using patients genetics to identify which drugs will kill them
-use gene chip which reads the degree to which the gene is active (on), vs (inactive) in the patients cancer
-each square identifies a particular gene, it will light up when gene expression of their DNA is in
-human genome project allowed this to happen
Genomics
scientific filed that sequences, interprets and compares whole genomes
(genome: complete DNA sequence)
-provides a list of genes present in the organism
functional genomics
examines when those genes are expressed and how their products interact. study of how all the products of a genome interact
human genome project
February 2001
-took 13 years to complete and cost 2.7 billion
-completed by International Genome Sequencing Consortium
when nutrients are suboptimal for cell growth and division, the cell cycle arrests (halts). At which cell cycle checkpoint would you predict this initial arrest to occur?
At the G1 checkpoint
The MPF protein complex turns itself off by:
Activating a process that breaks down cyclin
When a nucleotide substitution creates a STOP codon, it is
called ___________
A nonsense mutation
How are complete genomes sequenced?
may use a whole-genome shotgun sequencing approach:
-Genome is broken up into sets of overlapping fragments that are sequenced
-These sequences are then put in order
2 discoveries from human genome project:
- Genes for microRNAs are more common than previously thought. -miRNAs (microRNAs) are short non-coding RNAs that regulate gene expression post-transcriptionally.
- Many sequences are Transcripts of Unknown Function (TUFs) because their role in the cell is unknown.
* Significant insights have come from comparing the human genome to that of other species
Scientists do not know the function of more than:
a third of the genes found in the human genome.
Why do Humans have so Few Genes?
(alternative splicing hypothesis)
Eukaryotic genomes of organisms with complex morphology & behaviour * do not appear to have large numbers of genes.
Before the human genome was sequenced, scientists expected that humans would have 100,000 genes.
* actual sequence revealed that we have only about 21,000* genes. (way less)
The alternative-splicing hypothesis: proposes that eukaryotes do not require large numbers of distinct genes.
* Recall that alternative splicing creates different proteins from the same gene.
Comparison of Chimpanzee & Human genes
98.8% similar
* Most protein-coding sequences are similar, differences found in the regulatory sequences.
* Differences in how the protein coding genes are regulated may be responsible for the phenotypic differences seen in chimpanzees & humans.
Dr Tallulah Andrews
worked at the World Famous Wellcome Sanger Institute
-completed her undergrad studies
in Computational Biology/ Bioinformatics in the Biology Department McMaster University
research has focused on the analysis
of single-cell RNASeq data.
Researchers utilize tools created by advances in genomics to increase our understanding and knowledge of cancer
Can compare gene expression in normal vs cancerous cells.
* Human Genome Project has revealed common sets of genes that are mutated in cancerous cells. >120 distinct mutations may be involved
The complete genome sequences of cancerous & noncancerous cells from the same person identified over 600 mutations in the cancerous cells
Genome Canada
coordinates regional, national, and international genome projects in Canada & provides much of their funding
Since 2000, it has overseen 127 projects & >$1.8 billion in grants with most projects studying health issues.
(For example, how genetic makeup affects the response to pharmaceutical drugs. A drug may be beneficial for one person, inconsequential for a second person, & toxic for a third person.)
Laser Beam Experiment
Setup:
-fluorescent dye (chromosomes- blue, kineticore - yellow)
-anaphase: laser beam is photobleaching out fluorescent dye, compare length of microtubules from both sides, do they change with time?
prediction: photobleached section will still be visible as chromosomes begin to move
results: photobleached section remained visible, but the distance between chromosomes and photobleached section lessened.
Johnson & Rao’s Cell Fusion Experiments
not about sequence/order of cell cycle, just where cell cycle control enzymes are operating
-fuse 2 cells that are in diff phases (ex:mitotic and interphase cell)
result: DNA condenses in the interphase cell, start of M phase (means something is floating, something influenced the mitotic cell)
Markert & Masui’s Microinjection Experiments
2 cells in interphase (1 with M-phase cytoplasm, other w/interphase cytoplasm)
result: M-phase cell got spindle formation
conclusion of Johnson & Rao’s Cell Fusion and Markert & Masui’s Microinjection experiments
M-Phase Cytoplasm contains regulatory molecules that induces M phase in interphase cells
cancer cells:
nuclei: large, variably shaped
size/shape: variation
arrangement: disorganized
features: loss of normal features
many dividing cells
cancer cells divide faster than normal
cells
* uncontrolled cell division
* cell cycle is not regulated
* cell cycle checkpoints are disrupted
cancer cells exhibit excessive cell division
usually continue dividing well beyond a single layer, forming a clump of overlapping cells.
Cancer cells do not exhibit anchorage dependence. (normal cells anchor to dish surfaces and divide)
Cancer cells do not display density dependent inhibition (normal cells stop dividing once they form a single layer)
Stages of cancer
1: tumour is still small and has not grown outside the organ which it started in
2: larger than stage one but has not spread to nearby tissues
3: tumour is large and has spread to nearby tissues and lymph nodes
4: tumour has spread through the blood or lymphatic system to a distant site in the body
Chemotherapy Drugs
Vincristine (from rosy periwinkle)
Taxol (from Pacific Yew tree’s bark)
-are anti-microtubule agents.
- inhibit spindle fiber formation, which results disruption of mitosis
Steve Jobs
-Pim 1 Biomarker for pancreatic cancer
-Afinitor Drug
-Pancreatic cancer immunotherapy
CAR-T cell therapy for cancer
- collect white blood cells (t cells)
- each patients cells are assigned a number
- cells are frozen and grown in the lab
- the cells identify antigens
- cells infused into patient and binds to cancer cells to kill them
harnesses the potential of the individual’s own immune system to target & destroy cancer cells
All blood cells are derived from
self-renewing hematopoietic stem
cells that reside in bone marrow
erythrocyte (RBC) production
8 stages
-first 7 stages take place in the bone marrow
-Reticulocyte is then released into the bloodstream where it matures over 1-2 days into a functional erythrocyte (red blood cell)
do red blood cells have a nucleus?
do not have a nucleus, so cannot control cell growth and repair
-old and ruptured cells are replaced with new cells every 120 day
What controls how many red blood cells we make?
Oxygen levels regulate rate of erythropoiesis
The kidneys will mostly release the hormone erythropoietin (EPO) when blood oxygen levels are low and tissue oxygen demands are high
each red blood cell has:
-about 270 million hemoglobin molecules
Each hemoglobin molecule has 4, iron- containing heme groups (1 per subunit). Each one can bind oxygen
Inheritance of red blood cell type
We inherit our blood type from our parents
-Blood type is expressed from a single gene on chromosome 9 (ABO gene)
-The gene codes for a protein- glycosyltransferase,which modifies the carbohydrate content of the antigens on RBCs
antigens
The cell membranes of your red blood cells will have unique sugar-based markers
-specific to your blood cell type
(People with type A blood have type A
antigens on the surface of their RBCs)
(People with type AB blood have both type A and type B antigens on the surface of their RBCs)
(People with type O blood have no blood typing antigens on the surface of their RBCs -this make them universal donors)
If there is a mismatch during blood transfusion…
the immune system of the recipient will recognize these cells as “foreign” and will generate antibodies to the donor red blood cell antigen
These antibodies will bind the donor blood RBCs and clots (agglutination) will occur
..Because type O blood has no RBC surface antigens, they are not detected as foreign by the immune system
We determine red blood cell types
through “blood typing”
STEP 1: Take a glass slide with wells
STEP 2: Add blood sample
STEP 3: Add antibodies against
different antigens into the wells
if a person is type A: antibody against type B will show up, opposite for ppl with Type B
if a person is type AB: no antibody against A and B will showup
if a person is type O: antibody against both A and B will show up
who gives blood to who?
O to all (can only receive from O)
A to A and AB (receive from O,A)
B to B and AB (receive from B,O)
Ab only to AB (Ab can receive from all)
Dr. Stephen Withers & colleagues
University of British Columbia
Mucins (sugars that are attached to our cells such as the the glycoprotein) are very similar in structure to the sugar- based antigens that coat red blood cells
These enzymes can be added to donated blood to cut the antigens from any blood type and convert it to O-type
Some steps remain before we can use this innovation in medicine:
* This enzyme needs to be tested in animals and then in a human body on a larger scale
* We need to be sure it is safe in the human body and it will not lead to any complications in individuals that receive a blood transfusion that has this enzyme added
