2) Conditional alleles are commonly used by geneticists to control when (tempora
ID: 188000 • Letter: 2
Question
2) Conditional alleles are commonly used by geneticists to control when (temporal) and where (spatial) specific traits are expressed. In this question, you should outline a strategy utilizing the genetic tools that we have discussed during class. It might be simpler to draw a diagram of the engineered loci. Include all the components that are required for the experiment to be successful. A) What strategy could you use to switch off gene expression in liver cells? B) Why is a conditional allele valuable for scientists when studying gene function? C) What strategy could you use to switch on gene expression in liver cells in an adult mouse?Explanation / Answer
To switch off gene expression, the following methods can be used.
1. We can design an artificial miRNA/siRNA against the target gene. This transgene can be cloned under an inducible promoter if we want to repress gene function in a time frame.
2. We can use Cre-Lox system to knock-out a gene.
3. We can also employ antisense RNA technology.
Use of conditional alleles:
Conditional alleles are invaluable tools in genetic studies. Many of the genes in an organism are essential genes i.e, their loss function is embryonic lethal. So, the function of such genes can not be determined by traditional mutational analysis. So, scientists perform genetic screens to identify conditional alleles. For example, temperature sensitive alleles grow only in permissive temperature. They behave as WT cells at the permissive temperature. But, they exhibit mutant phenotype at the restrictive temperature. So, scientists allow the model organism to cross the early developmental stages at the permissive temperature and shift them to restrictive temperature at later developmental stages to observe their effect.
A gene function can be induced at two levels (Conditional induction = Gene can be expressed at desired time frame).
1. Transcriptional activation: Ethanol-inducible system
In this, the transgene is cloned under a promoter which is sensitive to ethanol. So, when the organism ingests ethanol, the gene is turned on.
2. Protein translocation: DEX-GR inducible system: This is applicable to transcription factors. As we know that transcription factors are functional in the nucleus but they are synthesized in the cytosol (Translation occurs in the cytosol in eukaryotes). So, a transgene is cloned with a GR tag (Protein fusion). When the protein gets translated, it will be sequestered by the HSP proteins in the cytoplasm (HSP binds to GR). When we add DEX to the system, GR binds to DEX relieving sequestration. Now, the protein is free to translocate into the nucleus and perform its function.
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