Transcript Objective - Central Magnet School

Single Nucleotide
Polymorphisms
Mrs. Stewart
Medical Interventions
Central Magnet School
Bell Work
• Why is Taq polymerase used in PCR
instead of human polymerase?
• Answer on your own paper
Objective
• Use laboratory techniques such as DNA
extraction, PCR, and restriction analysis to
identify single base pair differences in
DNA
• Explain how single base pair changes
called single nucleotide polymorphisms
(SNPs) can be identified through genetic
testing and often correlate to specific
diseases or traits.
Single Nucleotide
Polymorphisms
• The parts of the human genome that vary
by just a single nucleotide
• Abbreviated as SNPs
• Pronounced as “snips”
Effect of SNPs
• Non-coding DNA regions = no effect
• Genes that code for proteins = potentially
changing the protein produced
E
F
F
E
C
T
–Different phenotypes
–Disease/disorder
Remember:
• a change in DNA can lead to a change in
a protein
• If the protein plays a role in keeping you
healthy, serious consequences may occur.
Using a Single Nucleotide
Polymorphism to Predict Bitter
Tasting Ability
Lab Overview
Gene of Interest
• TAS2R38
• On chromosome #7
• Ability to taste “bitter”
What is your phenotype?
NonTaster
Weak
Taster
Strong
Taster
Step 1: Isolate a sample of your
DNA from your cheek cells
Step 2: Amplifying the Gene of
Interest
• Using your DNA sample, you will amplify a
220 base pair region of the PTC gene
using PCR.
– Specific primers attach to either side of the target sequence
SNPs for TAS2R38 gene
• In this lab, you will investigate one of the
base pair changes or single nucleotide
polymorphisms (SNPs) that affects a
person’s ability to taste the chemical PTC.
Genetics Review – Question 1
• The inability to taste PTC is a recessive
trait.
• If a capital “T” is is used to designate the
dominant allele and a lowercase “t” is used
to designate the recessive allele, what is
the genotype of a “Nontaster”?
Answer
• A “Nontaster” carries two recessive alleles
and thus has the genotype “tt”.
Genetics Review – Question 2
• What are the possible genotypes for a
“Taster”?
Answer
• A “Taster” may be homozygous dominant
with a genotype of “TT” or heterozygous
with a genotype of “Tt”.
• In this lab, you will use the tools of
molecular biology to determine your
genotype for PTC tasting.
Step 3: Restriction Analysis
• Restriction enzymes, molecular scissors,
recognize specific DNA sequences and
cut the nucleotide strands.
• In this part of the experiment, you will use
a specific restriction enzyme, HaeIII, to
identify a SNP or base pair difference in
the amplified segment of the PTC tasting
gene.
HaeIII and TAS2R38 gene
• HaeIII restriction enzyme
– 5’ GGCC
– 3’ CCGG
5’ ---GG
3’ ---CC
CC--- 3’
GG--- 5’
• TAS2R38 gene variations
NONTASTER (tt)
TASTER (TT)
GGCGGGCACT
CCGCCCGTGA
GGCGGCCACT
CCGCCGGTGA
Step 4: Gel Electrophoresis
• Gel Electrophoresis separates DNA
fragments based on their molecular
weight.
• Once you have digested your DNA sample
with the restriction enzymes, run your
product on a gel to analyze your results.
What will gel results show?
Non-taster
Taster