Genetics Lab Final

Product Rule (the AND rule)

For independent events – one event does not influence the outcome of the second event

MULTIPLY the individual probabilities of each event

Sum Rule (the OR rule)

For mutually-exclusive events – not independent, and one event precludes the possibility of the other occurring

ADD the total probabilities together

Positive control group

Groups which receive a treatment with a known response

Negative control group

Groups which receive a treatment known to have no response

Pseudoreplication

Treating observations as independent when they are not, leading to an inflated sample size

Probability (P)

Cutoff of p ≤ .05 (5%) is a widely accepted standard used to determine statistical significance

Degrees of freedom (df)

(# of categories) – (1)

Male D. melanogaster

epandrium

sex combs on legs

Female D. melanogaster

ovipositor

Beer’s Law

A=εlc

A = absorbance

ε = molar absorptivity (constant)

l = path length of light (constant)

c = concentration

High concentration = High absorbance

Pure DNA sample

1.7-2.0 absorbance at 260nm/280nm

SDS

Detergent part of Lysis Buffer

Disrupts cell membranes and denatures proteins

EDTA

Metal chelation (e.g. binding Mg2+) part of Lysis Buffer

Inhibits DNA degrading enzymes such as DNase

NaCl

Salt part of Lysis Buffer (positively-charged) to create an ionic environment, further protecting negatively-charged DNA from nucleases

Tris-HC

pH buffering to physiological pH (8.0)

Part of Lysis Buffer

KOAc (potassium acetate)

Concentrated salt solution

K+ binds and precipitates out SDS (from the Lysis Buffer), SDS-bound proteins, and other debris

Cold temperatures reduce solubility and increase precipitation

Supernatant After 1st Separation by Centrifuge

Supernatant: contains DNA, and some salts/debris
Pellet: debris

Alcohol DNA Precipitation

After 1st seperation

Isopropanol: quick and dirty precipitate with maximum DNA

Then ethanol: should remove the last of the contaminants

DNA Extraction Steps

1. Cell Lysis

2. Protein Removal

3. DNA Precipitation

4. DNA Rehydration

Polymerase Chain Reaction (PCR) Steps

1. Denature: temp. is raised to melt DNA strands

2. Anneal: temp. is lowered to anneal primer to target strand

3. Extension: using heat-stable Taq DNA polymerase