Lecture 27

Question 1

Disrupting gene expression:

Answer 1

• RNA interference
• inject dsRNA into embryos for particular genes to reduce the levels of mRNA for those genes
• Take hybridised probes for certain genes and get to know the pattern of expression for specific genes

Question 2

RNAi knockdown can produce changed in body plan:

Answer 2

• These changes in body plan are consistent with the known function of knock down
• This is good evidence for
• Correlation for the pattern of gene expression with body plan
• Disruption of these genes results in predictable changes of the body plan

Question 3

Disruption of Hox gene function changes body plan:

Answer 3

• But there isn’t enough proof

Question 4

Cis-regulatory sequences:

Answer 4

• Can be anywhere in the genome (5’ end, 3’ end etc)
• Changes in DNA which change the expression of their target gene.
• Changes in the coding regions (exons) of DNA have lead to altered body plans

Question 5

Evo-Devo:

Answer 5

• Evolution/de-evolution

Question 6

Ubx expression in a particular body part determines what that segment will become. How?

Answer 6

• Switching on or off the genes that determine the characteristics consistent with what that segment should be

Question 7

What is regulatory change? What happens when mutations occur in the cis-regulatory region?

Answer 7

• They could be regulatory as they could impact the expression of downstream genes
• This could effect the body plan

Question 8

Artemia (brine shrimp) compared to drosophila:

Answer 8

• Common ancestor 400my ago
• Artemia has lots of legs
• There are many conserved regions and many divergent regions

Question 9

Ectopic expression:

Answer 9

• Expression of a gene where it is not supposed to be expressed

Question 10

What happens when ectopically expressed drosophila and artemia Ubx occurs in drosophila?

Answer 10

• DmUbx repressed the formation of legs
• There is a different between the Ubx proteins of dros and artemia
• AfUbx is functional in dros
• Repression of limb formation in dros with DmUbx, but not AfUbx

Question 11

Dll- distillus:

Answer 11

• Expressed in limb primordia
• Ectopic expression of DmUbx represses Dll expression, while AfUbx represses very weakly
• AfUbx is functional because it has the same affect on Antp expression as DmUbx

Question 12

What is the difference between these proteins?

Answer 12

• There is a series of QA repeats on the terminal end of the proteins
• Artemia has a different sequence in this region
• The difference in body plan is encoded here
• Limb-repression function occurs here and limbs are formed, not repressed
• Replacement of this domain with QA will have repressed limb formation

Question 13

Does this Artemia and drosophila provide proof?

Answer 13

• Kind of
• But we are making inferences about what is happening in Artemia based on what we know happens in Drosphila
• We need more genome sequence and need the capacity to replicate genomes

Question 14

CRISPR:

Answer 14

• Can allow genes in any species to be manipulated
  1. CAS9 (an RNA guided protein) binds a sequence of DNA matching it’s programmable guide RNA
  2. It generates a ds break
  3. Repair enzymes repair the gap in the DNA using random nucleotides OR a sequence of your choice
• This means you could insert a modified Ubx gene, totally replacing the Ubx gene of Artemia (need a guide RNA etc but still) so you could study Ubx function!

Question 15

Diversification of body plan - how?

Answer 15

• Same gene sets - differences in regulation in various species (cis-regulatory sequences
• Same gene sets - differences in coding region in various species