Flashcards in Lecture 27 Deck (15):
Disrupting gene expression:
- 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
RNAi knockdown can produce changed in body plan:
- 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
Disruption of Hox gene function changes body plan:
- But there isn't enough proof
- 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
Ubx expression in a particular body part determines what that segment will become. How?
- Switching on or off the genes that determine the characteristics consistent with what that segment should be
What is regulatory change? What happens when mutations occur in the cis-regulatory region?
- They could be regulatory as they could impact the expression of downstream genes
- This could effect the body plan
Artemia (brine shrimp) compared to drosophila:
- Common ancestor 400my ago
- Artemia has lots of legs
- There are many conserved regions and many divergent regions
- Expression of a gene where it is not supposed to be expressed
What happens when ectopically expressed drosophila and artemia Ubx occurs in drosophila?
- 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
- 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
What is the difference between these proteins?
- 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
Does this Artemia and drosophila provide proof?
- 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
- 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!