Molecular GeneticsChapter 10

Part 1 – DNA & DNA Synthesis

What is DNA? • Stands for Deoxyribonucleic Acid• It is an organic molecule• Was known to be a chemical in cells by the end of the

nineteenth century.• Contains hereditary instructions. It is the chemical

code for every trait. The protein “blueprint”.• Can be copied and passed from generation to

generation.• DNA and RNA are both nucleic acids.

o They consist of chemical units called nucleotides.o The nucleotides are joined by a sugar-phosphate backbone.

Hershey-Chase Experiment DNA and RNA Structure

The Structure of DNA1. Nucleotides (4 types differ in bases)

a. Phosphoric Acidb. Deoxyribose sugarc. Nitrogenous bases:

Adenine-ThymineGuanine-Cytosine

2. Ladder Shape 3. Double strand, helix twist

Ladder Shape (Sides & Rungs)

Sides:• Phosphoric Acid• Sugar• Phosphoric Acid

Rungs:• A-T• T-A• G-C• C-G

SugarAcidSugarAcidSugar

T-AG-CC-GA-T

The model of DNA is like a rope ladder twisted into a spiral.

Structure of DNA

The Discovery of the Double Helix James Watson and Francis Crick determined

that DNA is a double helix. Watson and Crick used X-ray crystallography data to reveal

the basic shape of DNA.

Rosalind Franklin collected the X-ray crystallography data.

DNA Double Helix

Chromosome DNA Code:Genes =

• Segments of DNA

• Code for a trait

HairColor

EyeColor

DNA Chromosome Code:

Triplets aka Codons

• Sets of 3 Nucleotides

• Code for Amino Acids, that make proteins, that ultimately code for a trait.

Acid AcidSugar-T-A-SugarAcid AcidSugar-G-C-SugarAcid AcidSugar-C-G-Sugar

DNA Replication

– Chromosomes double in late interphase - 2n to 4n

– When a cell or whole organism reproduces, a complete set of genetic instructions must pass from one generation to the next.

– Watson and Crick’s model for DNA suggested that DNA replicates by a template mechanism.

– . DNA Replication ReviewDNA Replication Overview

2N

2N

• Begins at specific sites on a double helix.

• Proceeds in both directions.

Origins of Replication

Leading Strand

Lagging Strand

DNA Replication

Steps for DNA Replication:1. DNA untwists

2. DNA unzips

3. Corresponding base pairs Line up In sets of 3 nucleotides

aka “triplets” or “codons”

4. DNA reforms

5. 2 strands twist into helix IdenticalStrand

Flip to back side of “Copying DNA”Which diagram happens 1st?

Check: When does DNA replication take place?

Copying DNA–Replication: An Exact Copy-Practice

RNA Ribonucleic Acid“DNA messenger & taxi”

II. RNA Structure & Function

A. What is RNA?1. Organic Molecule2. Stands for Ribonucleic Acid3. Nucleic Acid4. Three Types

1. mRNA= messenger2. tRNA= transfer3. rRNA= ribosomal

B. Where is RNA located?1. mRNA in

nucleus & cytoplasm

2. tRNA only in cytoplasm

3. rRNA in the ribosomes

mRNA

mRNA

tRNA

rRNA

What is RNA’s structure?• Acid• Sugar-Base• Acid• Sugar-Base• Acid• Sugar-Base• Acid• Sugar-Base

1. Nucleotides=a. Phosphoric Acidb. Ribose sugarc. Nitrogenous

Bases: Adenine-UracilGuanine-Cytocine

2. Single Strand3. No Twisted helix

Comparison of RNA & DNA:

• Acid• Sugar-Base• Acid• Sugar-Base• Acid• Sugar-Base• Acid• Sugar-Base

Acid Acid

Sugar-Base-Sugar

Acid Acid

Sugar-Base-Sugar

Acid Acid

Sugar-Base-Sugar

Acid Acid

RNA DNA Ladder

Deoxy-ribose

Ribose

Uracil Thymine

What are RNA’s functions:

1. mRNA=• Copies the DNA code• Deliveries message to

Ribosome

Why not send the original DNA code out?

• DNA might be damaged!

• mRNA components are reused

• To copy more messages

Original DNA

mRNA copy

RNA function continued2. tRNA:• in cytoplasm• Picks up an

amino acid• “Taxis” the aa to

the Ribosome protein factories

tRNA

Aminoacid

RNA function continued

3. rRNA:

• in ribosome “protein factories”

• Protein synthesis

III. Protein Synthesis

Assembling Proteins from the DNA Instructions

Protein Synthesis

Transcription:

1. mRNA is copied off of DNA

2. In nucleus

3. Steps:

• DNA untwists

• DNA unzips

• RNA codons line up Transcription

Transcription:mRNA has:

• Ribose sugar

• Uracil instead of thymine bases

• Nuclear membrane allows it to leave!

DNA Code mRNA

A

T

C

G

U

A

G

C

Transcription the DetailsThe “start transcribing” signal is a nucleotide sequence

called a promoter.– The first phase of transcription is initiation:

• RNA polymerase attaches to the promoter.

• RNA synthesis begins.

– The second phase of transcription is elongation:• The RNA grows longer.

– The third phase of transcription is termination:• RNA polymerase reaches a sequence of DNA bases called a

terminator.

Translation• Conversion of

the message (mRNA Code)

• Into a protein

• By the ribosome factories

Translation 1. mRNA arrives at the

Ribosome (rRNA & protein)

2. tRNA picks up an amino acid (aa)

3. tRNA delivers the aa to the ribosome

4. aa are assembled into polypeptide proteins U A G C

GC

U

A

mRNA code

tRNA taxi

Translation: Summary

I = base (U) III = Amino acids connected by a peptide bondII = mRNA IV = tRNA V = amino acid

Translation: Details - 3 Phases

1. Initiation: Brings together:• The mRNA

• The first amino acid with its attached tRNA

• The two subunits of the ribosome

Note: An mRNA molecule has a cap and tail that help it bind to the ribosome.

Translation Continued2. Elongation: Also consists of several steps

• Codon recognition - The anticodon of an incoming tRNA pairs with the mRNA codon.

• Peptide bond formation - The ribosome catalyzes bond formation between amino acids

• Translocation - A tRNA leaves the P site of the ribosome. The ribosome moves down the mRNA.

• Elongation continues until the ribosome reaches a stop codon.

Translation Continued2. Termination

• Stop codon is reached and translation ends.

Making Proteins

• How do you know what amino acids are being coded for?

Summary:DNA Replication:• Make duplicate

DNA

• In nucleus

• Copy the chromosomes

• For cell division

Protein Synthesis:1. Transcription:

– Make mRNA– From DNA– In nucleus

2. Translation:– Make protein– Off mRNA code– Using amino acids– In cytoplasm

Part II Genetic Changes

Mutations• Any change in the DNA sequence

• Mutations may result from• Internal Agents such as rrrors in:

– Replication

– Transcription

– Cell Division

• Physical or chemical agents called mutagens.– External Agents

Causes of Mutations

• Spontaneous Mutations

• Mutagens– Agent that can cause a change in DNA– Radiation (X-rays, cosmic rays, Ultraviolet,

nuclear)– Chemicals (Asbestos, benzene, Formaldahyde)– High temperatures

Chromosomal Alterations

• Sometimes parts of chromosomes are broken off and lost during Mitosis and Meiosis

• Break and rejoin improperly during crossing over

• More common in plants

• Zygote usually dies, or is sterile

Mutations in Reproductive Cells

• Sperm or egg mutated can show up in offspring

• New trait– Sometimes benefits

• Protein that doesn’t work– Structural / functional problems– May not survive

Mutations in Body Cells

• Not passed to offspring

• Cell divides, new cells also have mutation

• Some mutations affect cell division– Uncontrolled cell division = Cancer

Point Mutations

• A change of a single base pair in DNA

• THE DOG BIT THE CAT

• THE DOG BIT THE CAR

Frameshift Mutation• A single base is added or deleted

• Shifts the reading of sets of 3 (codons)

• THE DOG BIT THE CAT

• THE DOB ITT HEC AT

Figure 10.21

What type are these?

Repairing DNA• Enzymes proofread the DNA and replace

incorrect nucleotides.

• Specifically the enzymes and proteins involved in replication can repair the damage.

• Work well, but not perfect

• More exposure to a mutagen the less likely it will be repaired

– Although mutations are often harmful,• They are the source of the rich diversity of

genes in the living world.

• They contribute to the process of evolution by natural selection.

Evolution Connection

Applied Genetics & Gene RegulationChapters 11 & 12

Haven’t our ancestors been able

to change the plants and

animals around them for thousands of

years?How? & Why?

Bellwork

How/Why did our ancestors modify their plants and animals?

•cross purebreds to make hybrids…•cross hybrids to select for recessive traits…

What types of traits were sought after?

Before the mid-1940’s, did they know they were modifying DNA?

•What if there is not any natural variation in a species to manipulate by breeding?•Is it possible to artificially create mutations in organisms and then see if those mutations can be bred? What could we do to purposely cause mutations?

radiation chemicalsheat

What are some types of things scientists can do with DNA, today?

DNA fingerprintingIdentify paternityIdentify innocence/guilt

CloningConvince an adult cell to grow into a whole new organism…as if it were a zygote.

Genetic EngineeringTaking a gene from one organism and put it in another organism

DNA Fingerprinting– On November 22, 1983,

• A 15-year-old girl was raped and murdered on a quiet country lane.

• Three years later, another 15-year-old girl was raped and murdered.

– DNA fingerprinting of DNA samples from suspects and the crime scene

• Proved one man guilty and another man innocent.

– DNA technology has rapidly revolutionized the field of forensics.

Murder, Paternity, and Ancient DNA

– DNA fingerprinting • Has become a standard

criminology tool.

• Has been used to identify victims of the September 11, 2001, World Trade Center attack.

• Can be used in paternity cases.

– DNA fingerprinting is also used in evolutionary research

DNA Fingerprinting Techniques

1. The polymerase chain reaction (PCR)

• a technique by which any segment of DNA can be copied quickly and precisely.

• Through PCR, scientists can obtain enough DNA from even minute amounts of blood or other tissue to allow DNA fingerprinting.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings

3. Gel Electrophoresis• Can be used to separate the DNA fragments obtained from different sources.

– The DNA fragments are visualized as “bands” on the gel. The bands of different DNA samples can then be compared.

DNA Fingerprinting Techniques

DNA Fingerprinting Techniques 2. Short tandem repeats (STRs)

• Are repetitive sequences of DNA that are repeated various times in the genome.

• Scientists use STR analysis to compare the number of repeats between different samples of DNA. They can prove that two samples of DNA come from the same person.

• Once a set of DNA fragments is prepared, the next step in STR analysis is to determine the lengths of these fragments.

Cloning

– The animals in these pictures all share two unusual features.• They are all members of

endangered species, and they are all clones.

– Some scientists are now focusing their efforts on cloning members of endangered species.

– The use of cloning to repopulate endangered species holds tremendous promise.

Cloning at the Edge of Extinction

Cloning Plants and Animals The Genetic Potential of Cells

– Differentiated cells• All contain a complete genome.• Have the potential to express all of an organism’s genes.

– Differentiated plant cells• Have the ability to

develop into a whole new organism.

• The somatic cells of a single plant can be used to produce hundreds of thousands of clones.

Cloning of Animals

– Regeneration, a similar process,• Is the regrowth of lost body parts in animals.

– Reproductive Cloning in Animals– Nuclear transplantation

• Involves replacing nuclei of egg cells with nuclei from differentiated cells.

• Has been used to clone a variety of animals.

– Scottish researchers cloned the first mammal in 1997.• Dolly, the sheep, was the product of their work.

The procedure that produced Dolly is called reproductive cloning.

Cloning of Animals

– Other organisms have since been produced using this technique, some by the pharmaceutical industry.

Cloning of Animals

Therapeutic Cloning and Stem Cells– Therapeutic cloning

• Produces embryonic stem cells (ES cells).

– Embryonic stem cells• Can give rise to specific

types of differentiated cells.

– Adult stem cells• Generate replacements for

nondividing differentiated cells.

– Umbilical Cord Blood Banking• Are unlike embryonic stem cells,

because they are partway along the road to differentiation.

Human Therapeutic Cloning– In 2001, a biotechnology company announced that it

had cloned the first human embryo.

Genetic Engineering akaRecombinant DNA Technology

Why in the world would we want to put DNA from one organism into another organism?

If a person needs a protein, but does not have the correct gene (or if their gene for a particular protein is mutated), …….

“I have diabetes…I will die without insulin.”

…then… How could they get that needed protein?

…..What are some examples of this?

“I am a hemophiliac…how can I make my blood clot?”

1. Get the protein from other humans

Problems:We typically don’t make more of a protein than we need…&…sharing of bodily fluids

can lead to sharing diseases… example, before we tested blood for HIV many

hemophiliacs got HIV from blood transfusions!

2. Use a similar protein from other animals

Example: insulin from pigs

Problems:Sometimes the protein is close,

but not exact & can lead to rejection problems…the process can be slow and

expensive

3. Make artificial proteins in the laboratory from chemicals…

Problems:very expensive and time consuming…hard to make perfect… the process of translation in a cell is regulated so well that there are rarely problems…humans make lots of mistakes in comparison!!!

4. Get bacteria (or another organism) to make the human protein in large amounts.

REALLY FAST!This is the thing to do…it is, NOW,

easy…cheap…and very fast!This is called Recombinant DNA…

to take DNA from one organism and put it into another organism in order to make a needed

protein.

Recombinant DNA Technology– Recombinant DNA technology is a set of techniques

for combining genes from different sources into a single DNA molecule.

• An organism that carries recombinant DNA is called a genetically modified (GM) organism.

– Recombinant DNA technology is applied in the field of biotechnology.

• Biotechnology uses various organisms to perform practical tasks.

From Humulin to Genetically Modified Foods

– By transferring the gene for a desired protein product into a bacterium, proteins can be produced in large quantities.

– In 1982, the world’s first genetically engineered pharmaceutical product was produced.

• Humulin, human insulin, was produced by genetically modified bacteria.

– Humulin was the first recombinant DNA drug approved by the FDA.

Genetically Modified (GM) Foods– Today, DNA technology is quickly replacing

traditional plant-breeding programs.• In the United States today, roughly one-half of the corn

crop and over three-quarters of the soybean and cotton crops are genetically modified.

– Corn has been genetically modified to resist insect infestation.

• This corn has been damaged by the European corn borer.

– “Golden rice” has been genetically modified to contain beta-carotene.

• Our bodies use beta-carotene to make vitamin A.

Genetically Modified (GM) Foods

Farm Animals and “Pharm” Animals– While transgenic plants are used today as commercial

products, transgenic whole animals are currently only in the testing phase.

– These transgenic sheep carry a gene for a human blood protein.

• This protein may help in the treatment of cystic fibrosis.

– While transgenic animals are currently used to produce potentially useful proteins, none are yet found in our food supply.

– It is possible that DNA technology will eventually replace traditional animal breeding.

Farm Animals and “Pharm” Animals

Recombinant DNA Techniques

– Bacteria are the workhorses of modern biotechnology.

– To work with genes in the laboratory, biologists often use bacterial plasmids.

• Plasmids are small, circular DNA molecules that are separate from the much larger bacterial chromosome.

– Plasmids can easily incorporate foreign DNA.– Plasmids are readily taken up by bacterial cells.

• Plasmids then act as vectors, DNA carriers that move genes from one cell to another.

– Recombinant DNA techniques can help biologists produce large quantities of a desired protein.

Recombinant DNA Techniques

taking a piece of DNA and cutting it to have “sticky ends”, then cutting a plasmid to have sticky ends, next, sticking the DNA in the plasmid, and finally, putting the plasmid back into a bacterium.

We can Give DNA to Bacteria by…

“Sticky ends”

Restriction Enzyme

Restriction enzymes cut the DNA so it has “sticky ends”!

What would be sticky in DNA?

Restriction Enzymes

1. Cut pieces of human DNA (genes) that code for the protein that is needed.

2. Cut open bacterial plasmids.

3. Stick the human gene in the plasmids.

4. Prepare bacteria to accept the plasmids by chemically tearing small holes in the cell walls and cell membranes & freeze them so they don’t ooze out the holes!

5. Briefly heat the bacteria to make the holes swell and allow the plasmids to sneak inside the bacteria.

6. Cool & feed the bacteria so their cell walls and membranes can heal…

7. Warm and feed the bacteria so they can grow…and produce the human protein.

The Basic Steps of Bacterial Transformation

Genomics and Proteomics– Genomics is the science of studying whole genomes.

• The first targets of genomics were bacteria.

The Human Genome Project– In 1990, an international consortium of government-funded

researchers began the Human Genome Project.• The goal of the project was to sequence the human genome.

– Sequencing of the human genome presented a major challenge.• It is very large.• Only a small amount of our total DNA is contained in genes that code

for proteins.

– The Human Genome Project• Can help map specific disease genes such as Parkinson’s

disease.

– The Human Genome Project proceeded through several stages,

• During which preliminary maps were created and refined.

The Human Genome Project

Genome-Mapping Techniques– The whole-genome shotgun method

• Involves sequencing DNA fragments from an entire genome and reassembling them in a single stage.

The Genetic Basis of Cancer– In recent years, scientists have learned more about the genetics of cancer.

– As early as 1911, certain viruses were known to cause cancer.

– Cancer-causing viruses often carry specific genes called oncogenes.

– Proto-oncogenes• Are normal genes that can become oncogenes.• Are found in many animals.• Code for growth factors that stimulate cell division.

– For a proto-oncogene to become an oncogene, a mutation must occur in the cell’s DNA.

– Tumor-suppressor genes• Help prevent uncontrolled cell growth.

• May be mutated, and contribute to cancer.

Human Gene Therapy

– Human gene therapy is a recombinant DNA procedure that seeks to treat disease by altering the genes of the afflicted person.

• The mutant version of a gene is replaced or supplemented with a properly functioning one.

Safety and Ethical Issues– As soon as scientists realized the power of DNA technology,

they began to worry about potential dangers such as:• The creation of hazardous new pathogens

• The transfer of cancer genes into infectious bacteria and viruses

– Strict laboratory safety procedures have been designed to protect researchers from infection by engineered microbes.

• Procedures have also been designed to prevent microbes from accidentally leaving the laboratory.

The Controversy over Genetically Modified Foods

– GM strains account for a significant percentage of several agricultural crops in the United States.– Advocates of a cautious approach have some concerns:

• Crops carrying genes from other species might harm the environment.

• GM foods could be hazardous to human health.

• Transgenic plants might pass their genes to close relatives in nearby wild areas.

– Negotiators from 130 countries (including the United States) agreed on a Biosafety Protocol.

• The protocol requires exporters to identify GM organisms present in bulk food shipments.

– Several U.S. regulatory agencies evaluate biotechnology projects for potential risks:

• Department of Agriculture• Food and Drug Administration• Environmental Protection Agency• National Institutes of Health

Ethical Questions Raised by DNA Technology

– Should genetically engineered human growth hormone be used to stimulate growth in HGH-deficient children?

– Genetic engineering of gametes and zygotes has been accomplished in lab animals.

• Should we try to eliminate genetic defects in our children?

• Should we interfere with evolution in this way?

– Advances in genetic fingerprinting raise privacy issues.

– What about the information obtained in the Human Genome Project?

• How do we prevent genetic information from being used in a discriminatory manner?

Evolution Connection:Genomes Hold Clues to Evolution– Genome data has confirmed evolutionary

connections,• Such as between yeast cells and human cells.

– Comparisons of completed genome sequences strongly support the theory that there are three fundamental domains of life:

• Bacteria

• Archaea

• Eukaryotes