CB - Screening and treatment of IEMs

Question 1

How are Tissue/DNA samples obtained from a Fetus?

Answer 1

Amniocentesis (12+ weeks)

• Fetal skin cells in amniotic fluid
• Centrifuge and culture

Chorionic villus sampling (8-12 weeks)

• Chorion is embryonic
• Risk of contamination by maternal tissue damage

Question 2

How are Tissue/DNA samples obtained from a Newborn and Adult?

Answer 2

Newborn
* Skin prick

Adult
* Blood sample
* Skin biopsy

Question 3

What can be used to test for Base substitutions?

Answer 3

ARMS (Amplification refractory mutation system)

• Uses allele-specific oligonucleotides as primers to test for mutations

Uses 2 PCR reactions to detect wt andd mutant strains

Question 4

What is DNA sequencing used for?

Answer 4

Scanning genes for unknown mutations

Question 5

What are biochemical treatments of IEMs? (4)

Answer 5

1. Control of metabolite consumption (PKU)
2. Supply of missing metabolite (GSD)
3. Drugs (statins) (FH)
4. No treatment (Tay-Sachs)

Question 6

What is the strategy for recessive and dominant gene therapy?

Answer 6

Recessive- Add normal gene
Dominant- Switch off mutant gene

Question 7

What stages are involved in Step 1- Preparing the corrective nucleic acid? (3)

Answer 7

1. PCR up normal version of gene
2. Need carrier for gene + promoter
3. Inserted into plasmid vector

Question 8

What are advantages and disadvantages for in-vivo? (4)

Answer 8

Advantages

• Only requires an injection
• Easy to re-treat

Disadvantages

• Non-specific targeting
• Immune response to the vector

Question 9

What are advantages and disadvantages for ex-vivo? (6)

Answer 9

Advantages

• No immune response
• No off-target effects
• Integration of transgenic DNA into genome

Disadvantages

• May require surgery
• Some cells are challenging to culture in vitro
• Poor engraftment rate

Question 10

What type of lipid helps fusions with the plasma membrane?

Answer 10

Cationic lipids help fusion as they fuse with Anionic lipids in the membrane