Single Nucleotide Polymorphisms

Mrs. StewartMedical 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

–Different phenotypes–Disease/disorder

EFFECT

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?

Weak Taster

Non-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 5’ ---GG CC--- 3’– 3’ CCGG 3’ ---CC GG--- 5’

• TAS2R38 gene variations

NONTASTER (tt) TASTER (TT)

GGCGGGCACT GGCGGCCACT

CCGCCCGTGA 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