24 Questions
What is the significance of using the C2H2 zinc finger motif as a platform for designing DNA binding modules with novel specificities?
It has enabled the production of novel ZFPs against investigator-specified loci
What is the benefit of using ZFNs in gene targeting?
It allows for bona fide reverse genetics in diverse model organisms
What is the advantage of using ZFNs over traditional gene targeting methods?
It eliminates the risk of insertional mutagenesis events
What is the significance of allele engineering in somatic cell genetics?
It is enabled by ZFNs
What is the benefit of using current-generation ZFNs in gene-function studies?
It removes constraints on experimental design
What is the significance of tagging an endogenous gene with GFP?
It enables the simultaneous elimination of artefacts driven by variegated expression levels
What is the significance of using ZFNs in cell-based therapy?
It enables the production of novel ZFNs against investigator-specified loci
What is the significance of ZFNs in gene targeting?
They enable the production of novel ZFPs against investigator-specified loci
What is the primary outcome of using ZFNs to introduce small insertions and/or deletions at the site of the break?
Disruption of a target gene
What is the role of an investigator-designed homologous donor DNA in gene correction?
To provide a template for repair
What is the result of ZFPs interacting with neighbouring fingers and bases outside their proximal DNA triplet?
Complicated efforts to produce new ZFPs
What is the outcome of using ZFNs in an embryo, resulting in up to 10% of the progeny being mutated for the gene of interest?
Gene disruption
What is the purpose of modular design in making ZFPs with new sequence specificities?
To enable more selective binding
What is the result of duplication events, usually with a subsequent divergence of function?
Formation of functional paralogues with new sequence specificities
What is the role of ZFNs in gene correction, in the presence of an investigator-designed homologous donor DNA?
To repair a double-strand break
What is the characteristic of non-functional paralogues, also known as pseudogenes?
Lack of promoter or intron sequences
What is the primary goal of creating a point mutation in a gene?
To study gene function and/or model disease-causing mutations
What is the advantage of using isogenic settings in gene-function studies?
It provides a way to study structure–function relationships at native levels of expression
What is the term for the process of creating a new allele of a gene?
Allele engineering
What is the main challenge in genetically engineering human stem cells?
Their recalcitrance to genetic engineering
What is the term for the study of the function of a gene by creating a mutation in the gene?
Reverse genetics
What is the application of gene targeting in human ES and iPS cells?
Modelling of human tissues (diseased and normal) in basic science, drug discovery, and regenerative medicine
What is the advantage of using ZFNs in gene targeting?
They enable the creation of point mutations at a specific position in the target gene
What is the term for the creation of a new allele of a gene through the correction of a point mutation?
De novo creation of point mutations
Study Notes
Zinc Finger Proteins (ZFPs) and Zinc Finger Nucleases (ZFNs)
- ZFPs can be designed to bind specific DNA sequences through modular design, but this can be complicated by interactions between fingers and neighboring base pairs.
- ZFNs can introduce small insertions and/or deletions at the site of a break, allowing for gene disruption or correction.
- ZFNs can be used to create point mutations in a target gene, mimicking disease-causing mutations or disabling motifs crucial for function.
Methods for Producing Novel ZFPs
- Duplication events can create new ZFPs with novel specificities through subtle divergence of function.
- Alternative strategies for making ZFPs include modular design, which can be complicated by interactions between fingers.
Gene Correction and Addition
- ZFNs can be used to correct point mutations in a target gene, allowing for the study of gene function and disease-causing mutations.
- ZFNs can be used to create isogenic panels of mouse ES cells carrying defined alleles for an endogenous gene.
- Gene addition in human ES and iPS cells can be used to model human tissues and study structure-function relationships.
Applications of ZFNs
- ZFNs have been used to create point mutations in D.melanogaster, with up to 90% of treated animals yielding offspring with donor-specified alleles.
- ZFNs can be used to study gene function and disease-causing mutations in human stem cells.
- The application of ZFNs removes many constraints on experimental design, allowing for more precise and efficient gene editing.
This quiz covers the concepts of gene duplication, mutation, and genetic variation, including the process of gene duplication events and the potential outcomes.
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