Class 21 - Molecular Variation: How to Assess it How do we...

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Unformatted text preview: Molecular Variation: How to Assess it How do we identify variation at the molecular level? Single base changes Restriction Fragment Length Polymorphism Allele Specific Oligonucleotides (ASO) DNA Sequencing Gene chips Multiple base changes Microsatellite repeat variation Minisatellite repeat variation (VNTR) Chromosomal changes Karyotype analysis Cytotology 2 3 Microsatellites - Simple Tandem Repeat Polymorphisms (STRP) Microsatellites are simple repeated elements of 2,3 or 4 bases (CA n , CTG n , CTAG n ) Position in genome is the same from person to person May have 10 or more alleles in a population (CA 5 ,CA 6... etc) Allele = difference is the number of repeats CACA, CACACACA, CACACACACACA Thought to arise by slippage of polymerase and/or unequal sister chromatid exchange 17 PCR is used to detect microsatellites GO TO PCR ANIMATION 4 6 Microsatellite assay CACACACA GTGTGTGT CACACacacACA GTGTGtgtgTGT PCR amplify with unique primers across CA repeat; length N PCR other homologue with same oligos will be length N+4 7 Real results Note - multiple bands due to slippage Homozygote Homozygote Hetero Hetero Hetero Hetero Hetero Hetero Hetero Hetero Hetero Hetero Hetero Hetero Homozygote Hetero 8 Microsatellites are good and bad Good news Highly polymorphic Large number of markers Assay may be automated Bad news Assay sucks Expensive Density of markers could be better Must do one at a time more or less 20 9 Please answer the following question without talking to your neighbor I.I....
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Class 21 - Molecular Variation: How to Assess it How do we...

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