Lab 4 – The Cell Cycle in Onion Root Tips The Need for Cell Division • During development – One fertilized egg cell begins to divide – Develops into a mature multicellular organism • In adults – To repair damage – To replace worn out tissue • Daughter cell vs. mother cell are identical Have same number of chromosomes Stages of the Cell Cycle • Before a cell can divide: – Key cellular components must be duplicated • Proteins, membranes, and organelles i.e mitochonderia, ribosomes, lysosomes – Chromosomes (DNA) must be duplicated • Complete set of genetic information – Ensures that the new cells can survive • When mitosis is out of control, it can lead to cancer ** Cell Cycle For cell to divide it has to go through cell cycle Interphase: cell doubles everything. 90% of the cell cycle. C1, S ,G2 phase: G1, cell grow S, chromosome duplicaion(DNA synthesis) G2, completes preparation for cell division Cell division. 2 processes; 1- mitosis In mitosis, nucleus divide --→ 2 nuclei 2- cytokinesis. In cytokinesis, cytoplasm divide. G1 S (DNA synthesis) G2 DNA Replication Mitosis Starts at Anaphase Interphase Mitosis Prophase Prometaphase Metaphase Anaphase Telephase Animal cells Interphase • Most cells are in interphase carrying out their functions ➢ Making insulin ➢ Secreting saliva ➢ Transmitting nerve impulses nucleolus nucleus Cell wall Plant cells Two main phases: Mitosis cell division 1- Mitosis; production of 2 nuclei • The division of nucleus and chromosomes • Consists of five phases – Prophase, Prometaphase, Metaphase, Anaphase and Telophase 2- Cytokinesis: division of the cytoplasm, produce 2 cells Prophase • Chromosomes condense – easier to separate without getting tangled together • Nuclear membrane dissolves • Microtubules (spindle fibers) attach to kinetochores Metaphase • Chromosomes line up at the center of the cell • Helps ensure that each daughter cell will get 1 copy of each chromosome Centrioles in animal cells only Anaphase • Chromosomes separate at the kinetochores • Chromosomes move to the opposite sides Telophase • Spindle fibers disappear • Chromosomes start to decondense • Nuclear membrane starts to reform • Cytokinesis (cell division) occurs Plant cells: cell plate formation Animal cells: cleavage furrow/pinching Late anaphase prophaseEarly anaphase metaphase Interphase Nucleus Nucleolus 1 Prophase Chromatin condensing Chromosomes 2 Prometaphase 3 Metaphase Cell plate 4 Anaphase 5 Telophase 10 µm Lab 4: Karyotyping – Visualization of Human Chromosomes • Humans have 23 pairs (46 total) of chromosomes • Distinguished by: ➢ Length ➢ Position of centromere ➢ Staining pattern using special dyes Idiogram Metacentric Submetacentric Acrocentric • Metacentric: centromere medially situated • Submetacentric: one chromosome arm is somewhat shorter than the other. • Acrocentric: quite near one end of the chromosome • Metacentric: centromere medially situated • Submetacentric: one chromosome arm is somewhat shorter than the other. • Acrocentric: quite near one end of the chromosome • Telocentric: Terminally situated (human do not have) Giemsa Staining (Banding Pattern = G-Banding) Karyotyping Detects Chromosome Abnormalities: 1. Congenital Disorders (By Birth) • People born with extra or missing chromosomes or pieces of chromosomes • Most chromosome abnormalities lead to miscarriages • Some exceptions: ➢Down Syndrome – 3 copies of chromosome 21 ➢Cri du Chat Syndrome – loss of a piece of chromosome 5 ➢Turner Syndrome – loss of one X chromosome ➢Klinefelter Syndrome – one extra X chromosome (XXY) • Amniocentesis: sampling of amniotic fluid using a hollow needle inserted into the uterus to screen for developmental abnormalities in a fetus Down Syndrome: 3 Copies of Chromosome 21 • Most common chromosome abnormality in humans: 1:1,000 live births • Phenotype: short stature, muscle weakness, heart defects, characteristic facial features Down Syndrome: 3 Copies of Chromosome 21 Cri du Chat Syndrome: Loss of a Piece of Chromosome 5 • Very rare: 1:15,000 – 1:50,000 live births • Phenotype: high-pitched cry, low birth weight & slow growth, intellectual disabilities, characteristic facial features Cri du Chat Syndrome: Loss of a Piece of Chromosome 5 Turner Syndrome: Loss of One X Chromosome • Occurs in 1:2,000 live births • Phenotype: sterility, short & stocky stature Turner Syndrome: Loss of One X Chromosome Klinefelter Syndrome: One Extra X Chromosome (XXY) • Occurs in 1:500 – 1:1,000 newborn boys • Phenotype: abnormal body proportions, sterility, increased risk of breast cancer Klinefelter Syndrome: One Extra X Chromosome (XXY) Karyotyping Detects Chromosome Abnormalities: 2. Chromosome Abnormalities That Occur Later in Life • Occur in one cell or a small group of cells • Can lead to cancer: ➢Philadelphia chromosome chromosomes 9 and 22 translocation between ➢Causes chronic myelogenous leukemia (CML; 95% of cases have Philadelphia chromosome translocation) Philadelphia Chromosome Philadelphia Chromosome • The abnormality seen by Nowell & Hungerford on chromosome 22, now known as the Philadelphia Chromosome • Both scientists worked at institutes located in Philadelphia HeLa Cells • Cells derived from a tumor removed from a woman named Henrietta Lacks in 1951 • Because they are tumor cells, they do NOT have normal number of chromosomes 70-90 chromosomes with over 20 translocations (line or population) Karyotyping Procedure – 1 Steps Performed Ahead of Time (page 62) 1. HeLa cells arrested in Metaphase • Chromosomes are duplicated & condensed • Can see where the Centromere is • Chemical called Colchecine arrests cells in Metaphase 2. Cells are put in a hypotonic solution (more solutes inside the cell than outside) • Water enters the cells & they swell 3. Cells are fixed in Acetic Acid/Methanol to preserve their structure Karyotyping Procedure – 2 Today’s Lab Starting With Step 13 (p. 65 – 68) 1. Use a pipette to suspend cells by gently sucking them up & down in the pipette several times 2. “Splat” cells onto COLD microscope slide (place slides at 45˚ angle). Add 5 drops ONE DROP AT A TIME. 3. Blow on the slide (2-3 sec) to spread the cells 4. Allow slide to dry completely (incubator) 5. Follow staining procedure on p. 43 (steps 16 and 17) *** Wipe the bottom edge of slides between stains *** 6. Rinse slide in a beaker of water & allow to dry (incubator) 7. Add Permount to the coverslip, place onto the slide, & press gently 8. Examine slide under the microscope 9. Count chromosomes in 5 different cells Lab 4 (Mitosis) Report Worksheet (8 points) • # of cells in each phase of the cell cycle • Interphase (I), Prophase (P), Metaphase (M), Anaphase (A), and Telophase (T) • If you spotted any, mention a few words about cells undergoing cytokinesis • % of cells in each cell cycle phase (I, P, M, A, T) • This should be easy if you counted a total of 100 cells • Time spent in each cell cycle phase (I, P, M, A, T) • • Just multiply 24 hours (example of the length of cell cycle given in your lab manual; it can be different in the exam) with the percentage of the cells in each phase Express your results in hours (hr) and minutes (min) e.g., 10.4 hr = (10 + 0.4) hr = 10 hr and (0.4 hr x 60 min/hr) = 10 hr and 24 min Lab 4 (Karyotyping) Report Worksheet (4 points) • Please fill-in-the-blanks and short answer questions ➢ Work on your own ➢ Look up the answers to the questions in lab manual (glossary, background, procedures, tables, etc.) Biology 115 Lab Fall 2018 Lab 4 Worksheet Instructors: _____________________________________________________________________________________ I. Student Name: Group Members: _____________________________________________________________________________________ II. Personalized Title [0.5 pts] _____________________________________________________________________________________ III. Hypothesis [1.0 pts] 1. _____________________________________________________________________________________ IV. Introduction IVA. General Theory/Background [1.0 pts] IVB. Summary of Overall Approach [1.5 pts] Page 1 of 3 Biology 115 Lab Fall 2018 Lab 4 Worksheet Instructors: _____________________________________________________________________________________ V. Observations & Results VA. Data [1.5 pts] VB. Calculations [0.5 pts] Page 2 of 3 Biology 115 Lab Fall 2018 Lab 4 Worksheet Instructors: _____________________________________________________________________________________ VI. Discussion Questions 1. Define mitosis and cytokinesis. What are the results of each? [0.5 pt] 2. How does cytokinesis differ in animal and plant cells? [0.5 pt] 3. What important biological characteristics of life depend on mitotic cell division? [0.5 pt] _____________________________________________________________________________________ VII. Conclusion(s)/Inference(s): [0.5 pt] _____________________________________________________________________________________ Total Score: _____ / 8.00 (Includes _____ Bonus Points for ________________________________________________) Comments: ___________________________________________________________________________ _____________________________________________________________________________________ Page 3 of 3 Biology 115 Laboratory Fall 2018 Instructors: Full Name: _________________________________ Lab Section: _______________________________ Lab 4 Report Worksheet A. Fill in the blanks (0.5 pt esch): 1. ___________________________________ syndrome is characterized by a deletion of the short arm of Chromosome #5? 2. The structure within the cell _________________________. that contains 3. A “matched pair” of chromosomes, ________________________________. one the genetic from each material is parent, called are the called 4. An ________________________ is any chromosome other than the sex chromosomes. B. Fill out the following table as you view your chromosome spreads under the microscope (1.0 pt): Cell # Chromosomes # Metacentric # Submetacentric # Acrocentric # Telocentric 1 2 3 4 5 C. Short answer questions (0.5 pt each): 1. What was the purpose of this experiment? __________________________________________________________________________________ __________________________________________________________________________________ __________________________________________________________________________________ __________________________________________________________________________________ 2. Why do some cells have more or fewer than 46 chromosomes? __________________________________________________________________________________ __________________________________________________________________________________ __________________________________________________________________________________ __________________________________________________________________________________ __________________________________________________________________________________ __________________________________________________________________________________ __________________________________________________________________________________ Page 1 of 1 
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