DNA Analysis

Lesson 1.2PBS

What is DNA?

• Deoxyribonucleic Acid (DNA)– A double-stranded, helical nucleic acid molecule capable of

replicating and determining the inherited structure of a cell’s proteins.

• Human DNA is a unique code of over three billion base pairs that provides a genetic blueprint of an individual.– The fundamental building block of life.– Hereditary material found in all living organisms.– Found in nucleus of cells.

What is DNA?

• DNA is packaged as chromosomes, which each contain numerous genes, or segments of DNA sequence that code for traits.

• Chromosomes: any of the usually linear bodies in the cell nucleus that contain the genetic material.

• Gene: a discrete unit of hereditary information consisting of a specific nucleotide sequence in DNA (or RNA, in some viruses).

– Consists of thousands of genes that contain instructions for building different parts of the cell.

– Determines physical characteristics we have and some of the diseases we will develop.

DNA Structure

• DNA from all living organisms has the same basic structure – the differences are in the sequences of the nucleotides.

• Nucleotide– A building block of DNA,

consisting of a five-carbon sugar covalently bonded to a nitrogenous base and a phosphate group.

Pentose Sugar

• 5-Carbon sugar called deoxyribose.

• Found on outside of DNA molecule.

• Covalently bonded to phosphate.

Nitrogenous Bases• Adenine

– A component of nucleic acids, energy-carrying molecules such as ATP, and certain coenzymes. Chemically, it is a purine base.

• Cytosine– A component of nucleic acids that carries hereditary information in DNA and RNA

in cells. Chemically, it is a pyrimidine base.• Guanine

– A component of nucleic acids that carries hereditary information in DNA and RNA in cells. Chemically, it is a purine base.

• Thymine– A component of nucleic acid that carries hereditary information in DNA in cells.

Chemically, it is a pyrimidine base.

• Base pairs found on inside of DNA molecule.• The sequence of these bases determine an organism’s traits.

Phosphate Group

• Found on the outside of the molecule.

• Covalently bonded to deoxyribose sugar.

Double-Helix

• Helix: something spiral in form.– Antiparallel double helix.

• Chargaff’s Rules:– Adenine binds to Thymine

• Amount of A=T• 2 Hydrogen Bonds

– Guanine binds to Cytosine• Amount of G=C • 3 Hydrogen Bonds

How does DNA differ from person to person?

• The difference between different people’s DNA is only about 0.1%.

• The sequence of nucleotides differs from individual to individual creating a unique DNA profile for every person.– Only identical twins have the same DNA profile.

How do scientists isolate DNA in order to study it?

• DNA Extraction– Can be isolated from plant and animal cells.– Take advantage of unique chemical properties of the

DNA.– Cell Components:

• Cell walls/cellulose (PLANTS ONLY)- physical action to break down.

• Cell membranes/nuclear membranes- detergents/soaps break down.

• Proteins- meat tenderizer breaks down.• Ethanol removes contaminates like proteins and other cellular

components.

DNA Can Be Extracted From:

• Cells in:– Bone– Blood– Hair Root & Shaft– Semen– Saliva– Teeth– Urine & Feces– Fingernail Debris– Muscle Tissue

– Cigarette Butts– Postage Stamps– Envelope Sealing Flaps– Dandruff– Fingerprints (Touch DNA)– Personal Items: razor

blade, chewing gum, wrist watch, ear wax, toothbrush.

Challenges When Extracting

• Contamination• Degradation- heat, humidity, UV damage.• Mixed Samples (Will show multiple profiles)

How can tools of molecular biology be used to compare the DNA of two

individuals?• DNA Extraction• DNA Amplification- process called Polymerase

Chain Reaction (PCR) where multiple choices of the DNA sample are made.

• DNA Analysis– Restriction Enzymes– Gel Electrophoresis

What are restriction enzymes?

• Enzymes are proteins that speed up chemical reactions.

• Restriction Enzyme (Restriction Endonucleases):– A degradative enzyme that recognizes specific

nucleotide sequences and cuts up DNA.– Attaches to the DNA and reads the base sequence;

then breaks the covalent bonds between the sugar and phosphate at the restriction site and “cuts” the DNA into multiple pieces.

– Found in bacteria.

Restriction Enzymes: Molecular Scissors

• Cut DNA at specific sequences

• What kinds of bonds are broken when restriction enzymes cut?– Covalent bonds (within a

single strand) – Hydrogen bonds

(between strands) as a result of the strands coming apart

Hydrogen bond Covalent bond

What are restriction fragment length polymorphisms?

• Restriction Fragment Length Polymorphisms (RFLPs) – Differences in DNA sequence on homologous

chromosomes that can result in different patterns of restriction fragment lengths (DNA segments resulting from treatment with restriction enzymes).

– Different segments of DNA; measured in base pair length/number of base pairs.

– Analyzed using gel electrophoresis.

What is gel electrophoresis and how can the results of this technique be

interpreted?

• Since the sequence of DNA is to small to see, gel electrophoresis separates DNA fragments based on size and electrical charge.

• Measure their rate of movement through an electrical field in a gel.

Gel Electrophoresis

• DNA is loaded into an agarose (sugar) gel.• An electrical current is supplied to separate

the different size DNA fragments.• DNA has a negative charge (from the

phosphate groups) and is repelled by the negative side of the apparatus and attracted to the positive side.

• The DNA shows up as bands on the gel.

Smaller pieces of DNA move faster (further) because they have less frictional resistance in the agarose.

DNA Profiles or DNA Fingerprints

DNA Profiling Uses

• Can be used to:– Match suspect and crime scene DNA.– Match a baby with possible fathers.– Match bone DNA or body parts with a missing

person DNA.– Track genealogy.– Study bio-history (Anthropology & Archeology).– Diagnose disease (gene therapy, genetic

counseling, genetic conditions).