Positional cloning Flashcards
To do positional cloning, what do we have to know about the disease gene?
Approximate location in the genome
What are the 5 basic steps in positional cloning?
- Define the candidate region
- Obtain clones of all the DNA in that region
- Identify all the genes in the region
- Prioritize them for mutation screening
- Test candidate genes for mutations in affected people
How do you find the candidate region?
Use high density and high resolution linkage maps with lots of defined markers
Why do we want to use high resolution maps for positional cloning?
More reference points means we can more precisely narrow down where the disease gene is
What are unlinked loci?
Loci that assort independently from each other
What are non-syntenic unlinked loci?
Loci that assort independently and are on different chromosomes
What are syntenic unlinked loci?
Loci that assort independently and are located far apart on the same chromosome
What are linked loci?
Nearby loci on the same chromosome that generate fewer recombinant gametes than parental gametes due to a decreased likelihood of crossing over
What is the maximum likelihood of crossing over between two loci?
50%. They’re unlinked once they reach 50%
What is informative meiosis?
Meiosis that lets us determine unambiguously which gametes are parental and which are recombinant
What are two things we have to know in order for meiosis to be informative?
Heterozygosity and phase
What is phase?
The location of each allele. Knowing which ones are together on the same chromosome and which are on different homologous chromosomes
What are alleles in coupling?
On the same homologous chromosome
What are alleles in repulsion?
On different homologous chromosomes
What is haplotype?
The combo of closely linked DNA sequences on one chromosome that are frequently inherited together
How do we determine phase from a pedigree?
Need the third generation and at least one of the parents in the second generation is a heterozygote for the loci of interest. The third generation lets us unambiguously determine phase
Can we definitively determine the distance between markers using a pedigree?
No, human families are small. We can tell if two markers are close together, but can’t tell how far apart they are exactly
What are 4 traits that make a desirable genetic marker?
- Co-dominant alleles
- Polymorphic loci
- Well distributed throughout the genome
- Easy to detect
What are the 4 things we typically use as genetic markers?
RFLPs, minisatellites, microsatellites, and SNPs
Does any marker fit the criteria of an ideal marker perfectly?
No, none of them are perfect. But are still good for different reasons
