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Using a Single Nucleotide Polymorphism
to Predict Bitter Tasting Ability
Bridges 2014
Taste
 Taste is mediated by specialized cells that communicate with
several brain regions through direct connections to sensory
neurons
 Taste perception is a two-step process:
 A taste molecule binds to a specific receptor on the surface of a
taste cell
 Taste cells generate a nervous impulse, which is interpreted by
the brain
Bitter Taste Discovery
The inability to taste phenylthiocarbamide (PTC) is a recessive trait
that varies in the human population.
PTC Taste Regulated by SNP
 The gene for the PTC taste receptor, TAS2R38, was
identified in 2003
 Sequencing identified three nucleotide positions that vary
within the human population—each variable position is
termed a single nucleotide polymorphism (SNP)
 One specific combination of the three SNPs, termed a
haplotype, correlates most strongly with tasting ability
 SNPs also predict adverse responses to PROZAC® and
Paxil®
Today’s Activities
 Figure out YOUR genotype
 Extract DNA from your cheek cells
 Boil
 Chelex resin, which binds contaminating metal ions
 PCR amplify a portion of the TAS2R38 gene
 Digest the PCR product with restriction enzyme HaeIII
 Recognition sequence includes one of the SNPs
 One allele is cut by the enzyme, and one is not
 Produces a restriction fragment length polymorphism (RFLP)
 Gel visualize the RFLP
 Compare genotype to phenotype
DNA Extraction
DNA Extraction
 Boiling
 Lyse cell (break apart/open up)
 Chelex
 Binds contaminating metal ions-major inhibitor of PCR
 Spin
 Precipitate out debris
Supernatant
Supernatant
Supernatant
Pellet
Polymerase Chain Reaction
 Create many copies of a SPECIFIC portion of DNA
 Requires:
 Ready-To-Go PCR bead
 Taq Polymerase
 Buffer
 MgCl2
 Nucleotides- A, G, C, T
 Primer/Loading dye mix (on ice!)
 Primers to target Portion of the TAS2R38 gene
 Loading dye for gel
 Template DNA- cheek DNA from Part I
Thermocycling
Exponential Growth of Template DNA
PCR
Restriction Enzyme Digest
 Enzyme- protein that carries out chemical reactions
 Restriction enzymes cut DNA at a SPECIFIC nucleotide
sequence called a restriction site
 The HaeIII restriction enzyme uses GGCC recognition site
 It only digests the TAS2R38 gene at this site
 Remember the TAS2R38 contains single nucleotide
polymorphisms
 You may be GGCC and/or GGGC
 The HaeIII may or may not digest your DNA
Restriction Enzyme Digest
Gel Analysis