Exam #5: Molecular Medicine I Flashcards Preview

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Flashcards in Exam #5: Molecular Medicine I Deck (47)
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How are antibodies used in western blotting?

Technique that uses specific antibodies to indicate the molecular weight of a protein

-Tells if the protein is present or not
- Used to quantify the amount present
- Shift in banding can identify a modification of the protein in question


How are antibodies used to treat cancer?

- Herceptin was the first monoclonal antibody approved for treatment of metastatic breast cancer
-inhibits ErbB2/ HER2/Neu receptor tyrosine kinase located on cell surface


What will DNA sequencing show & not show?

- Will show if the person is homozygous or heterozygous
- Will NOT show which chromosome a mutation is on
- Will NOT show a deletion or duplication


Comparative Genome Hybridization (CGH)

1) Obtain ss-DNA from patient & control
2) Label both with different dye
3) Mix & hybridize to metaphase chromosomes
4) Uneven labeling (color) indicates an insertion or deletion (variation in copy number)


What are the steps in PCR?

1) Denature DNA (want ss-DNA)
2) Synthesize oligonucleotides that are complimentary to the ends of the DNA fragment to be studied (primers)
3) Primers anneal to ss-DNA
4) Polymerase synthesizes complimentary strand
5) Repeat


Real-time PCR

Allows for quantification of amounts of nucleic acids

- Measures amplified DNA after each cycle
- Determined how many cycles are required to pass a threshold
-Template amount can be quantified by the number of cycles required to reach threshold
- More cycles= less template & vice versa


Reverse-Transcriptase PCR

Used for RNA analysis

- Reverse transcription of RNA to DNA by reverse transcriptase
- Makes cDNA
- cDNA serves as the template for the PCR reaction


Warfarin example of pharmacogenetics

- Oral anticoagulant that inhibits vitamin K epoxide reductase
- polymorphism in p450 (CYP2C9) causes slow metabolism & higher sensitivity
- polymorphism in vitamin K epoxide reductase (VKORC1) that leads to warfarin tolerance


What cells produce antibodies?



What is a sandwich ELISA routinely used for?

The detection of antigen or peptide in human serum

1) Parathyroid Hormone
2) HbA1c
3) CRP


What is an indirect ELISA routinely used for?

Diagnosis of autoimmune disease

- RA
- Also, HIV


What are the steps in a western blot?

1) Separate protein extracts by electrophoresis
2) Transfer proteins to membrane
3) Add enzyme-linked specific antibody
4) Antibodies bind antigen (proteins)
5) Detect label in band pattern


Gene Therapy

Insertion of a functional copy of a gene into mutant cells


What is the biggest obstacle to gene therapy?



What viruses are used as vectors in gene therapy?

1) Retrovirus
2) Adenovirus
3) Adeno-associated virus (No known disease)
4) HSV-1


What are the conflicting properties of a viral vector?

- Virus should spread aggressively through the body; however, if TOO AGGRESSIVE, can trigger sepsis
- Integration into the host genome, which can lead to cancer (leukemia in clinical trials)


What are the four non-viral vectors in gene therapy?

1) Liposomes= lipid vesicle w/ DNA payload
2) Naked DNA
3) Complexed DNA
4) Artificial Chromosomes