Lecture 8 - Genetic Variation

Question 1

Define gene silencing

Answer 1

To determine the role of a gene you can knockout (completely switch off) or knockdown (reduce) expression of a gene

Question 2

Name some major approaches of gene silencing

Answer 2

Antisense (knockdown)
Short Interference RNAs (knockdown)
CRISPR (knockout)

Question 3

Explain gene knockdown

Answer 3

Use Watson and crick binding (A-T(U)) and C-G) by RNA/DNA molecules to guide proteins that modify the target gene or mRNA
Recruit a protein which knockdown the gene expression

Question 4

Define siRNA-mediated RNA interference

Answer 4

RNA interference is mediated by the RNA interference silencing complex (RISC) siRNAs direct RISC to specific mRNAs, determined by base pairing of the target mRNA and siRNA
Target coding regions of the mRNA
Design one strand that is complementary (antisense) to target the mRNA
Once bound Ago 2 catalyses cleavage of the mRNA and will be degraded

Question 5

Explain antisense knockdown

Answer 5

Targets the levels of the mRNA in the cell
1) single-stranded RNA/DNA of 15-30 nucleotides that is complementary (antisense)
2) use predicted RNA structure programmes to target areas of mRNA that are single-stranded and accessible
3) DNA/RNA complex, RNaseH recognises a DNA sequence bound to the RNA sequence, cleaves mRNA becomes unstable and gets chopped up by ex-nucleases
4) Eventually leads to knockdown of the protein

Question 6

What is CRIPSR?

Answer 6

Used to target many genes at once advantage to diseases not caused by single mutation
CRISPR/Cas9 is a bacterial defence mechanism against virus infection
can be used to cleave human DNA and produce gene knockdown