Week 9

Question 1

advantages of viral vector systems (3)

Answer 1

high efficiency and expression
virus coating DNA.
can be broad or selective

Question 2

Adenoassociated virus are used for? Why?

Answer 2

Therapy in humans. It does not integrate.

Question 3

What virus are used in research?

Answer 3

Retrovirus and lentivirus

Question 4

disadvantages of viral vector systems?

Answer 4

Activation strategies complex due to high security requirements
Therapeutic concerns (can insert themselves into oncogene, can have allergic reaction...)

Question 5

Two types of genes in virus?

Answer 5

Early: involved in replication. Promote entry of host into S-phase.
Late: structural. Capsid proteins and envelope proteins.

Question 6

Replication-competent addition what is it? what is a disadvantage?

Answer 6

Add gene of interest.

Limited by packaging size of virus

Question 7

Replication-competent replacement what is it? what is a disadvantage?

Answer 7

Replaces virus gene w gene of interest.

It is helper-independent.

Question 8

Helper-dependent vectors

Answer 8

Cannot replicate on its own and needs function supplied in trans.

Question 9

In vitro cloning

Answer 9

Cut with nucleases and put in transgene w ligase

Question 10

In vivo cloning

Answer 10

Insertion through homologous recombination

Question 11

How to make Simian Virus SV40 safe?

Answer 11

Take out T-antigen complex, which is what signals for unwinding of DNA. Needs COS cell line to replicate now.

Question 12

Adenovirus Genome changes to make it safe.

Answer 12

Packaging signal (psi) needed for the virus to package. Can make virus w no psi, or w psi and missing late genes to prevent unwanted gene replication.

Question 13

Adenovirus genome advantages (3)

Answer 13

Broad host range among vertebrates
Dividing and non-dividing cells can be infected
Low pathogenicity

Question 14

Adenovirus genome disadvantages

Answer 14

Previous exposure can lead to reduced efficacy
Inflammation
Leaky expression

Question 15

Lytic cycle basics?

Answer 15

Phage DNA into bacterium, host cell transcribes and translates, phage particles assemble, lyse host cell and is released.

Question 16

Lysogenic cycle basics

Answer 16

Phage accidentally acquires fragments of host DNA and transduction occurs upon infection of new host (integration of fragments into host)

Question 17

Adeno-associated viruses (AAV) advantages

Answer 17

Different tropisms allows for programming of genome anyway wanted.
Change of capsid gene structure allows to target different organs.

Question 18

What are 4 challenges for gene therapy?

Answer 18

Maximise impact but minimise risks
Strategies that work on animal models may not work in humans
Research is time consuming and difficult (many steps)

Question 19

How is ex gene therapy usually made?

Answer 19

Isolate haematopoietic stem cells and then put then back in the organism. Has limited cell divisions.

Question 20

Gammaretroviral vector characteristics

Answer 20

Replicate in host cells via reverse transcription

BUT can integrate into proto-oncogenes

Question 21

CAR-T cell therapy

Answer 21

take blood, target cancer, culture, put receptor cell that can select against cancer and then put it back. Can bind to cancer and kill it.

Question 22

Advantages of CAR-T cell therapy

Answer 22

ex vivo, which is ideal, and can be used with CRISPR CAS9, better than virus.