Genetic Molecular Techiniques - Part V - RNAi, Human Disease
Genetic Molecular Techiniques - Part V - RNAi, Human Disease
Genetic Molecular Techiniques - Part V - RNAi, Human Disease
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The Solution: dsRNA directs mRNA Degradation!<br />
input dsRNA<br />
in vitro transcription<br />
of dsRNA<br />
RNA interference (<strong>RNAi</strong>)<br />
processing<br />
RNase<br />
~22nt siRNAs<br />
5’ 3’<br />
3’ 5’<br />
target<br />
recognition<br />
mRNA<br />
5’ 3’<br />
3’ 5’<br />
degradation<br />
Fig. 9-43a<br />
injection of dsRNA<br />
degradation of target<br />
mRNA in progeny - “KO”<br />
-specific<br />
-almost any mRNA can be<br />
targeted<br />
-defense against RNA viruses<br />
-works in human cells<br />
-high throughput screening<br />
of gene function with <strong>RNAi</strong><br />
“libraries”<br />
Analysis of Gene Function by <strong>RNAi</strong> KO<br />
1. Synthesize dsRNA in vitro<br />
2. Inject parent worm with dsRNA<br />
3. Test effect on mRNA expression by in situ<br />
4. What is the phenotype?<br />
Fig. 9-43b<br />
mex-3 mRNA present mex-3 mRNA absent<br />
9.6 Identifying and Locating <strong>Human</strong> <strong>Disease</strong> Genes<br />
KEY CONCEPTS:<br />
1. Patterns of inheritance: Autosomal dom.,<br />
Autosomal recessive, X-linked rec.<br />
2. Recombinant genotypes by crossing over<br />
3. Mapping - cosegregation with markers<br />
4. <strong>Molecular</strong> markers - polymorphisms<br />
5. RFLPs, SNPs, SSRs<br />
6. Expression analysis, DNA sequencing<br />
7. <strong>Genetic</strong> heterogeneity and polygenic traits
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