Module 9

Question 1

DOP-PCR (WGS method)

Answer 1

• Degenerate-oligonucleotide-primed PCR
• Good for degraded samples but no increase in sensitivity
• Low-stringency stage = 3’ anneals to 106 sites then adjacent hexamer binds

Question 2

MDA (WGS method)

Answer 2

• Multiple displacement amplification
• More robust due to higher fidelity polymerase
• Amplify low amount DNA by annealing random hexamers primers then copied with ϕ29 polymerase
• Best for over 2000 bases and not good for degraded samples

Question 3

RCA (WGS method)

Answer 3

• Rolling circle amplification
• DNA must be circularized: T4 DNA ligase, CircLigase II (no concatemers), MIPs
• Random oligonucleotide primers = circle means never ending template
• Exonuclease resistant random primers + ϕ29 polymerase: proofreading so no degradation

Question 4

Capture/genomic partitioning

Answer 4

• Enriches DNA for regions of interest
• Method 1 = streptavidin-coated magnetic beads bind to regions of interest
• Method 2 = biotinyl probe added then hybridized with circular genomic ssDNA template

Question 5

MIPs

Answer 5

Ss oligonucleotide with 2 complementary terminal regions flanking SNP of interest

Question 6

Host factors

Answer 6

• Like linking scratch to fingernails, wounds to bullets, and bites to teeth
• Immunoglobulin response