PV92 PCR Informatics
- 1. 11/06/11 Any SINE of Chromosome 16? PV92 PCR/Informatics ( S hort In terspersed Repetitive E lement)
- 2. Think of your entire genome as one very fat book of biological data. 11/06/11 Like this...
- 3. 100 pages total: - 95 pages of nonsense - 5 pages of useful information
- 4. Only 5 pages of your genetic book contain useful information!
- 5. 11/06/11 Today we are going to look in a non-coding region of your chromosome #16 But remember: You got one chromosome #16 from Mom And one chromosome #16 from Dad Mom thinks hers is better.
- 6. Alu repeats: our targets Classified as SINEs ( S hort In terspersed Repetitive E lement) Mobilized by an RNA polymerase - derived intermediate (retroposition) Approx. 500,000 Alu copies per haploid genome, representing about 5% of the genome Named for the Alu I restriction site within the element 11/06/11
- 7. Evolutionary Significance of PV92 Alu Inserts Highly conserved Inserted in the last 1,000,000 years Genotypes (+/+, +/ - , - / - ) Used in population genetics, paternity analysis, and forensics 11/06/11
- 8. 11/06/11 PCR Results The PV92 Alu is dimorphic so there are two possible PCR products: 641 bp 941 bp No insertion: 641 bp 300 bp Alu insert 641 bp Alu insertion: 941 bp 5 ’ 3 ’ Alu Amplified Region
- 9. 11/06/11 Actual Alu PCR Results 941 bp 641 bp - +/- + + - +/-
- 10. Amplify Alu insert from representative sample population Calculate the expected allelic and genotypic frequencies 11/06/11 To estimate frequency of Alu within a population:
- 11. 11/06/11 InstaGene Extraction Cell membrane Nuclear membrane Heat disrupts membranes InstaGene matrix binds released cellular Mg ++ Genomic DNA Mg ++ Mg ++ Mg ++ Mg ++ Mg ++ Mg ++