BCH 5413 Exam 1-Questions With
100% Verified Answers.
Northern Blot Controls - ✔✔1. equal loading of all RNA in lanes
2. Quality of RNA
3. Intact RNA was actually transferred
4. Internal mRNA standards to control equal loading and transfer
5. Positive and negative controls
Western Blot Controls - ✔✔1. Housekeeping protein that doesn't change with treatment
2. Quality of Sample
3. Other possible interactions with antibody
4. positive and negative experimental controls
Positive control - ✔✔makes sure reagents worked
correctly Northern: probe for something you know will show
a change
Western: load a lane withe recombinant protein you know will give signal
Negative control - ✔✔tests to see if NO signal when required reagent is
missing Western: using secondary antibody only
Equal loading controls - ✔✔did you really put the same amount of sample into each
lane? probe for GAPDH or actin
Cross-reactions - ✔✔Could one of your reagents be detecting something else as
well? use BLAST (NCIB website)
, Thermes Aquatic - ✔✔heat stable polymerase (Taq polymerase)
PCR Denaturing - ✔✔At 95 degrees C for 30 seconds
PCR annealing primers - ✔✔between 30-50 degree C for 30 seconds
PCR extending primers - ✔✔at 72 degree C for 1 minute
PCR reaction components - ✔✔1. Template DNA
2. Primers (forward and reverse)
3. dNTP (G, A, C, T)
4. Taq polymerase
5. Buffer + magnesium
6. Water
Uses of PCR - ✔✔1. Cloning
2. Genotyping (Triplex PCR)
3. Analyze DNA methylation patterns (Bisulfite PCR)
4. Characterize DNA and protein interactions (footprinting)
5. amplify rare footprint (Ligation-mediated PCR)
DNA Footprinting - ✔✔used to analyze sequence-specific DNA-protein interactions
•DNA footprinting occurs when digestion of a specific region of DNA is prevented by binding
of a protein. The protection can be either to digestion by DNases, or to chemical modification.
The second process gives greater resolution.
***The footprint represents the region of
100% Verified Answers.
Northern Blot Controls - ✔✔1. equal loading of all RNA in lanes
2. Quality of RNA
3. Intact RNA was actually transferred
4. Internal mRNA standards to control equal loading and transfer
5. Positive and negative controls
Western Blot Controls - ✔✔1. Housekeeping protein that doesn't change with treatment
2. Quality of Sample
3. Other possible interactions with antibody
4. positive and negative experimental controls
Positive control - ✔✔makes sure reagents worked
correctly Northern: probe for something you know will show
a change
Western: load a lane withe recombinant protein you know will give signal
Negative control - ✔✔tests to see if NO signal when required reagent is
missing Western: using secondary antibody only
Equal loading controls - ✔✔did you really put the same amount of sample into each
lane? probe for GAPDH or actin
Cross-reactions - ✔✔Could one of your reagents be detecting something else as
well? use BLAST (NCIB website)
, Thermes Aquatic - ✔✔heat stable polymerase (Taq polymerase)
PCR Denaturing - ✔✔At 95 degrees C for 30 seconds
PCR annealing primers - ✔✔between 30-50 degree C for 30 seconds
PCR extending primers - ✔✔at 72 degree C for 1 minute
PCR reaction components - ✔✔1. Template DNA
2. Primers (forward and reverse)
3. dNTP (G, A, C, T)
4. Taq polymerase
5. Buffer + magnesium
6. Water
Uses of PCR - ✔✔1. Cloning
2. Genotyping (Triplex PCR)
3. Analyze DNA methylation patterns (Bisulfite PCR)
4. Characterize DNA and protein interactions (footprinting)
5. amplify rare footprint (Ligation-mediated PCR)
DNA Footprinting - ✔✔used to analyze sequence-specific DNA-protein interactions
•DNA footprinting occurs when digestion of a specific region of DNA is prevented by binding
of a protein. The protection can be either to digestion by DNases, or to chemical modification.
The second process gives greater resolution.
***The footprint represents the region of
