Microscopy Lab Techniques

Question 1

List and explain each Bacterial Growth Curve phases

Answer 1

1. Lag phase: Adaptation prior to cell division
2. Exponential (log) phase: Rapid doubling
3. Stationary phase: Growth rate = death rate
4. Death phase: Decline due to lack of food/other variable

Question 2

Density centrifugation

Answer 2

1 cycle where organelles are separated by density into layers

Question 3

List the most dense to least dense organelles in the density centrifugation layers

Answer 3

Nuclei
Mitochondria/chloroplast
ER fragments
Ribosomes

Question 4

Blood centrifugation

Answer 4

Type of density
centrifugation with 3 layers: Plasma, Buffy coat & Erythrocytes

Question 5

List blood centrifugation layers & what composes the first 2 layers

Answer 5

1. Plasma: Clear fluid
2. Buffy coat: Leukocytes & Platelets
3. Erythrocytes

Question 6

What is Polymerase Chain Reaction (PCR)

Answer 6

Amplify the amount of a DNA sample
3 steps: Denaturation, Primer annealing, Elongation

Question 7

List & explain PCR layers

Answer 7

Denaturation (~95 °C): heating separates DNA into single strands
Primer annealing (~65 °C): DNA primers hybridize with single strands
Elongation (~70 °C): nucleotides are added to the 3’ end of DNA using Taq polymerase

Question 8

Gel electrophoresis?

Answer 8

Separates DNA fragments by charge and size
An electric field is applied to agarose gel
Restriction enzymes
*Smaller fragments** travel further to the bottom of the gel
Sodium dodecyl sulfate detergent used

Question 9

In gel electrophoresis, where is the neg. cathode and pos. cathode?

Answer 9

Top = negative cathode
Bottom = positive anode

Question 10

What is the blotting acronym?

Answer 10

SNOW DROP
Southern blotting = DNA

Northern blotting = RNA

nOthing = nOthing

Western blotting = Protein

Question 11

What does the Western blotting primary & secondary antibodies

Answer 11

Treated with primary antibody (binds to target protein) & secondary antibody (attached to indicator and binds to primary antibody)

Question 12

Pulse chase experiments phases explained

Answer 12

Pulse phase: amino acids are radioactively labeled & incorporated into proteins
Chase phase: prevents radioactively labeled protein production

Staining helps track proteins throughout the cell

Question 13

Chromatography

Answer 13

Separating components of a heterogeneous sample using differential solubility
- 2 Phases: Mobile & Stationary

Question 14

List & explain chromatography phases

Answer 14

Mobile: sample is dissolved in the solvent
Stationary: mobile phase climbs up the apparatus & ascend to different heights

Question 15

SDS-PAGE

Answer 15

Separates proteins rather than DNA
Similar to gel electro

Question 16

DNA microarray

Answer 16

Fluorescent probes to identify which genes are being expressed in an organism.