Topic 1b

Question 1

LUCA

Answer 1

last universal common ancestor

Question 2

What domain(s) do we find prokaryotes in?

Answer 2

• archae and bacteria

Question 3

Cell Culture

Answer 3

• plated on the surface of a solid substance bathed in a nutrient medium
• normal contact-inhibited, anchorage-dependant human cells are attatched to the surface of a coated culture dish as a monolayer
• anchorage-independant tumor cells attach to the surface of a coated culture dish and grow as foci

Question 4

Normal Cells

Answer 4

• anchorage-dependant; usually attatched to the extracellular matrix and experience highly regulated growth
• dependant on growth factors provided in serum
• contact-inhibited; stop dividing because of declining growth factors or contact
• form monolayer at confluence
  -have a finite lifespan to their Hayflick limit; normal HDF ~50-100 doublings

Question 5

Transformed (malignant) Cell

Answer 5

• anchorage-independant (can grow in suspension)
• growth factor independant (do not need serum, but can grow better in it)
• not contact-inhibited
• can grow beyond a confluent monolayer (form foci, or multiple layers)
• grow indefinitely

Question 6

Preparation of a Primary Culture

Answer 6

1. use mechanical or enzymatic disruption to break down a tissue into individual cells
2. place cells in a suitable environment

Question 7

First Human Cell Culture

Answer 7

• Henrietta Lacks cervical cancer sample collected by Dr. George Gey and Margaret Gey
• HeLa cells; henrietta passed away two years later, and family did not know for decades that her cells were alive

Question 8

Cryopreservation

Answer 8

Subjecting organisms to low temperatures - cryobiology: studying the effect of the above

Question 9

Advantages of Cryopreservation

Answer 9

Biological backups: ensure cells of interest will always be available in the future (research, medical, commercial uses)
- allows for repetitions of the same experiment
- store cells for additional work at different stages at differentiation or after a variety of experimental treatments
- “suspended animation” stops biological time when cells are stored at < -130ºC

Question 10

Cryoprotective Agents

Answer 10

Help to prevent danage caused by the freezing process.
- initial discovery by Polge, Smith, and Parkes (1949) using chicken spermatozoa with glycerol as the cryoprotectant
- most common cryoprotectants for eukaryotic cells DMSO (dimethylsulphoxide) at 5-15% v/v or glycerol at 5-20% (v/v) or more

Question 11

Appearance of Cells (4 phases)

Answer 11

1. Abioitic synthesis of simple organic compounds (ie. amino acids, nitrogenous bases)
2. Abiotic polymerization of these monomers into macromolecules (ie. protiens, DNA)
3. Generation of a macromolecule capable of storing genetic information and replication
4. Encapsulation of this molecule in a membrane, forming a primitive cell

Question 12

RNA Subunits as First Informational Molecule

Answer 12

• RNA monomer subunits as precursors to DNA monomer subunits
• riboenzymes (RNAs folding into shapes and serving as enzymes) can synthesize short pieces of RNA in presence of nucleotides

Question 13

Liposomes

Answer 13

hollow, membrane-bound vesicles of differing sizes that form when lipids are mixed with water
- can carry out metabolic reactions in presence of enzymes and their substrates
- primordially may have trapped RNA, forming protocells

Question 14

Prokaryotes

Answer 14

• bacteria and archaea
• non-nucleated, no membrane-bound organelles (DNA is folded into a compact structure called a nucleoid)
• small (1-5 um)
• divide via binary fision
• circular DNA, associates with protiens
• 70s ribosomes

Question 15

Eukaryotes

Answer 15

• large (10-100um)
• divide via mitosis/meiosis
• 80S ribosomes

Question 16

Archaea

Answer 16

one of the three domains of life (bacteria, archaea, eukarya)
- encompasses many species that live in extreme habitats on earth and have diverse metabolic strategies
- believe to be descended from a common ancestor of eukaryotes, long after it diverged from bacteria
- much more similar to eukaryotes than bacteria in DNA replication, transcription, RNA processing, and initiation of protien synthesis
- unique features are ribosomal RNA and membrane phospholipids
- DNA is tyically circular and complexed with moderate amounts of protiens that resemble eukaryotic histone

Question 17

Methanogens

Answer 17

• type of archaea
• obtain energy from hydrogen while converting carbon dioxide to methane

Question 18

Halophile

Answer 18

Grow in extremely salty environments
-archaea

Question 19

Macidophiles

Answer 19

Thrive in acidic hot springs where pH is as low as 2 and temperature can exceed 100 degrees celsuis.
- archaea

Question 20

Limitations on Cell Size

Answer 20

1. the requirement for an adequate surface area/volume ratio
2. rates at which molecules diffuse
3. need to maintain adequate local concentrations of substances involved in cellular processes

Question 21

Surface Area/ Volume Ratio Limitation

Answer 21

as a cell increases in size, surface area does not keep pace with volume, and so necessary exchange of substances cannot occur

Question 22

Rate of Diffusion Limitation

Answer 22

• rate of diffusion decreases as size of molecule increase; size of cell is limited by diffusion rates of molecules it contains
• can be bypassed by active transport for ions, macromolecules, etc
• another bypass: cytoplasmic streaming/cyclosis; active movement/mixing of cytoplasmic components

Question 23

Need for Adequate Concentrations of Reactants and Catalysts Limitation

Answer 23

• amount of reactants and catalysts must increase proportionally with cell size
• can be bypassed via compartmentalization of activities (localizing certain processes witin specific regions of the cell that need their products) in organelles

Question 24

Cyanobacteria

Answer 24

• posess extensive internal membrane systems on which photosynthesis is carried out
• one of few types of prokaryotic cells which have internal membrane systems

Question 25

Chromatin

Answer 25

• multiple linear molecules of DNA that are intracitely complexed with great amounts of protiens known as histones
• eukaryotic cells

Question 26

clonal culture

Answer 26

cell culture with a known genetic lineage

Question 27

axenic culture

Answer 27

only one species is grown in a given
culture

Question 28

cells are typically grown either…..

Answer 28

-suspended in a liquid nutrient medium (nutrient broth of unicellular organisms, suspension of transformed animal cells)
-suspended in semi-solid nutrient medium (ie. anchorage independant tumour cells in sloppy agar)
-plated on the surface of a semi-solid nutrient medium (ie. colonies or lawn of bacteria on 1-2% agar)

Question 29

cells are typically grown either…..

Answer 29

-suspended in a liquid nutrient medium (nutrient broth of unicellular organisms, suspension of transformed animal cells)
-suspended in semi-solid nutrient medium (ie. anchorage independant tumour cells in sloppy agar)
-plated on the surface of a semi-solid nutrient medium (ie. colonies or lawn of bacteria on 1-2% agar)

Question 30

basal medium components

Answer 30

L-amino acids
vitamins: function as components of coenzymes
salts
serum (dialyzed for cell nutrition, whole for stock cultures): the exogenous source of growth factors
required by normal cells
water
misc:
- glucose
- anitiboitics (ie. penicillin, streptomycin): reduce the risk of bacterial
contamination in this very rich medium
-phenol red

Question 31

current cyropreservation techniques

Answer 31

-controlled ice nucleation
-temp. dropped to -80c at a rate of 1c/min