LECTURE - DNA Sequencing

Question 1

absolute quantification

Answer 1

• to measure the # of molecules of the gene target in a group of samples
• method std curve

Question 2

relative quantification

Answer 2

• to measure relative change in the quantity of a gene between two or more samples
• std curve method or comparative Ct

Question 3

functions of Std Curves

Answer 3

• take efficiency into account
• convert Ct’s to quantity units
• test dynamic range

Question 4

different sequencing methods

Answer 4

• Sanger sequencing (dideoxynucleotide chain termination)
• pyrosequencing
• bisulfite DNA sequence
• sequencing by synthesis (next gen)
  > Illumina and Ion Torrent

Question 5

Sanger sequencing Workflow

Answer 5

• extraction
• PCR
• sample preparation
• Sanger Sequencing
• Capillary electrophoresis
• analysis

Question 6

DNA sequencing is used in the lab for a variety of reasons:

Answer 6

• detecting mutations
• typing microorganisms
• identifying human haplotypes
• designating polymorphisms