The Human Genome And Gene Mapping
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The Human Genome And Gene Mapping
Readings
Jorde, Carey, Bamshad & White: Medical Genetics, 3rd edition, C.V. Mosby Publishing, 2005.
- Structure of the Human Genome Chapter 2, 17-22
- Detection of variation at the DNA level Chapter 3, 43-55
- Gene Mapping and cloning Chapter 8, 160-189
Objectives
The student will:
- Understand the different types of genetic variation.
- Be able to distinguish between genetic mapping and physical mapping.
- Understand the basic concept of genetic linkage analysis
- Understand different gene mapping techniques .
The Human Genome Project
- Goal is/was determination of the complete sequence of the
human genome
- genes which define our phenotype
- genes whose alteration leads to disease
- Characterization of single gene disorders, multigene disorders
- 3 x 109 bp (<5% encodes genes)
- 24 different chromosomes
- First draft of the sequence of human genome published in 2000
Mapping
- Genome maps are detailed constructs of the order and/or position of genetic markers and DNA sequence.
- Maps are of two types:
- Genetic maps - use recombination frequencies of markers at meiosis
- Physical maps - identify exact location of DNA sequence in the genome
Human Genome Mapping
- Individual chromosomes isolated, and used to prepare genomic DNA libraries in yeast artificial chromosomes
- Unusual world-wide collaboration
- New technology of robotic production lines (PCR tests)
- Information shared on web (900 printed pages)
Genetic Markers
- Freely recombine during meiosis unless they are physically close together
- Linked markers usually inherited as a set
Genetic Maps
Genetic maps dependent on linkage analysis and recombination frequency:
- Units of scale
- Human = centimorgan (cM)
- 1 cM = recombination fraction of 1/100 meioses
- Equivalent to 0.7 - 1 Mb of DNA
Definitive genetic map
- Assembled by Genethon laboratory (Paris)
- Used microsatellite markers
Map Your Disease
- Find a large family with a clearly defined disease
- Using PCR check with 400 primers pairs, at about 11 cM intervals
- Automation
- Markers used in mapping
- Restriction fragment polymorphisms (RFLP)
- Minisatellites
- Microsatellites
Linkage Calculation
- LOD score = log of the odds score = statistical estimate of whether two loci are likely to lie near each other on a chromosome
- If they are they are likely to be inherited together
- LOD score > 3 generally taken to indicate that the two loci are close (1 to 1,000)
- Determine recombination frequencies - can range from 0% (no recombination) to 50% (crossing over in half of meioses)
- 50% recombination frequency would be observed from two markers on different chromosomes
- Not calculated by hand...
Probabilities
- Probability found by multiplying together the probabilities of each event happening independently
- For instance: I need to be at work by 8 AM. For this to happen I have to set my alarm clock, I have to be lucky with traffic, and I have to find a parking space
- Probability (work at 8) = Probability (set alarm) x Probability (no traffic) x Probability (parking space)
Physical Maps
- Plot the actual location of DNA sequences on chromosomes
- Units of scale = base pairs
- Clone maps
- Radiation hybrid maps - radiation breaks chromosomes
- Fluorescent in situ hybridization (FISH)
- Karyotype changes
Useful Terms
- Genetic markers - provide the landmarks to plot positions within the genomic landscape
- May be associated with phenotypic characteristics such as inherited diseases
- Many not associated yet with observable phenotype = DNA sequence polymorphisms
Array Technology
- Possible because of new technology
- Allows better resolution than comparative genomic hybridization (CGH) on chromosomes
Array Analysis
- Uses include gene expression, CGH, SNP analysis, protein analysis
- Differences lie in material on array (DNA, cDNA, oligonucleotides, antibodies, proteins)
- Choice of targets on array
- Sensitivity - minimum region
- Mosaics may not be detected
- DNA - can do multiple tests in one experiment
- Can compare multiple different individuals at multiple different targets
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