From Gene To You

A Look at Chapters 14-21

DNA as Genetic Material

Deoxyribonucleic Acid

Hershey & Chase

Hypothesized the structure

Structure of DNA

Nitrogen BaseAmounts vary from species to

speciesNitrogen RingsPurines are double ring bases

Adenine, Guanine

Structure of DNA con’t

Pyrimidines are single ring basesThymine, Cytosine

A-T uses two hydrogen bondsG-C uses three hydrogen bonds

5 Carbon SugarCalled Deoxyribose

Structure of DNA con’t

Phosphate GroupsBond with Sugar to form

backbone

The Double Helix

Watson & Crick used x-ray testing, previous studies to come up with double helix model

Sugar-Phosphate backbone with Nitrogen Base rungs

10 Layers (rungs) per turn

The Double Helix con’t

Base Pairing allows for equal amounts on each strand

Anti-parallel: one strand is oriented 3’ to 5’, the other 5’ to 3’

Dictates complements, buts allows for infinite variation

DNA Organization

Packaged with proteins to form matrix called chromatinCoiled around Histones to form Nucleosomes.In non-dividing cell:Euchromatin- Heterochromatin

The Structure of RNA

Ribonucleic Acid

Sugar is Ribose, not deoxy

Uracil replaces Thymine as the compliment of Adenine

Single stranded

Protein Synthesis

A little lesson in logic………Traits are the end products of

metabolic processes regulated by enzymes or created by polypeptides

DNA codes for all enzymesDNA codes for all polypeptides

Protein Synthesis con’t

Therefore…..the DNA needs to be read and somehow changed to be useful to the cell and organism

Process is called Protein Synthesis

Protein Synthesis con’t

Three steps:TranscriptionSynthesis of RNA using DNA as a

Template

RNA processingModifies the RNA to make it

functional

Protein Synthesis con’t

TranslationProteins are synthesized

according to genetic message of sequential codons along mRNA

Three types of RNA complete the process

Protein synthesis

Types of RNA

mRNA (messenger) is the template for Amino Acids to form the polypeptideCodon: triplete group of

nucleotides that codes for specific AA’s

64 codons = 20 AA’s

Types of RNA

tRNA (transfer) transports AA’s to proper place on the mRNA templateAnticodon is the compliment of the

mRNA codon (mirror)

rRNA (ribosomal) builds the ribosomes

Protein Synthesis- Transcription

Nucleotide sequence transcribed from DNA to compliment mRNA

mRNA carries code to Ribosome

Initiation:RNA Polymerase unzips DNA

Transcription con’t

Elongation:RNA Polymerase unzips and

assembles mRNA using DNA template

Termination:RNA Polymerase reaches

AAAAAAA, (stop Nucleotides)

transcription

Protein Synthesis- RNA Processing

Code is proofread and modified before leaving NucleusMakes a functional mRNA Eliminate Introns so specific proteins can be made by ExonsIntrons may be key to cell variation

Rna Processing

Protein Synthesis- Translation

tRNA is interpreter between the base sequence mRNA and the AA sequence in Polypeptide

Proteins coordinate the pairing of tRNA anticodons to mRNA codons

Translation con’t

InitiationTakes the mRNA and attaches to

initiator tRNA and 2 ribosomal subunits to assemble ribosome

ElongationAdd AA’s 1 by 1 to Initial AA’s

Translation con’t

Codon recognition used to assembe the peptide bonds (hydrogen bonds) to form the polypeptide

TermationUAA, UAG, UGA are stop codonsCompleted Polypeptide, last tRNA,

and Ribosomal subunits released

translation

The Genetics of Viruses

Cell Specific

A Nuclei Acid surrounded by a Protein Coat (Capsid)

A membrane coats some viral Capsids called an Envelop

Can kill cells, produce toxins

Viruses con’t

Some partially damage cells that eventually regenerate (Flu)

Some permanently damage cells that do not reproduce (Polio)

Viral Life Cycle-General

Infect host cell with viral Genome

Co-Opt Host’s Resources to:Replicate Viral GenomeManufacture Capsid Proteins

Assembling of new viral Nucleic Acid for next generation

Viral Life Cycle

Somewhat specific Life Cycles

Lytic Viral Life Cycle is where replication results in death or lysis of host cell

Are considered Virulent

Lytic Cycle:

Lytic Cycle

Penetration-using enzymes to destroy host cell DNA, and to replicate viral DNA

Transcribes viral DNA into RNA

Translates RNA to proteins

Assemble proteins and DNA into new Virus

Lytic Cycle con’t

New viral proteins erupt from host cell, destroying the host cell

Off to new cell to begin anew

Lytic Cycle

Lysogenic Cycle

Viruses co-exist with host by incorporating viral genome into host genome

Called Temperate Viruses because either Lytic or Lysogenic

Lysogenic Cycle con’t

Penetration- like Lytic cycle, but does not destroy host DNAInserts by Genetic Recombination (Crossing Over) into host genome, called a Provirus (Prophage)Inactive there until trigger, goes Lytic

Lysogenic Cycle

RNA Viral Life Cycles

Sometimes- viral RNA is used directly as mRNA

Retrovirus- a double stranded RNA genome, use negative strand as mRNA template

Transcribe DNA from viral mRNA

RNA Cycles con’t

Use Reverse Transcriptase to make DNA compliment

DNA then used to either make mRNA (Lytic) or chills (Lysogenic)

Viral Life Cycle

Viroids

Viroids are plant pathogens

Simpler than viruses,and smaller

Small naked circular RNA

Do not encode protein, but do replicate in host plant cells

Disrupt metabolism

Prions

Protein pathogens that cause degenerative brain diseaseDefective versions of normal proteinsCannot replicate, but hypothesis is they convert normal protein to prion protein, chain reaction

The Genetics of Bacteria

Bacteria contain 1 singular, circular DNA with no histones

Located in Nucleoid Region of Cell

Reproduce by binary fission

Contain Plasmids

Genetics of Bacteria con’t

Plasmids are short, circular, double stranded DNAShort life span facilitates evolutionary adaptation to environmentGenetic Recombination produces new strains, separate from fission

Bacterial Genetic Recombination

TransformationBacteria absord DNA from

surroundingsSpecial Proteins on surface

recognize and import DNA from closely related species

Genetic Recombination con’t

TransductionGene is transferred by a virus

BacteriophageDuring Lytic Cycle, incorporates

Bacterial DNA, carries to new cell when it incorporated into new Genome

Gentic Recombination con’t

ConjugationTransfer DNA between two bacterial

that are temporarily joinedTube is called Pilus (F-plasmids)R-plasmids give resistance to

antibiotics, make resistant strains

Regulation of Gene Expression

Activation of specific genes at specific times

Most often tested example…the bacterium E. coli (loves your digestive tract, especially your large intestine!)

Begins with Operons, sequences of DNA that direct biosynthetic pathways

The Operon-Four Components

Regulatory Gene produces a repressor protein (prevents gene expression by blocking RNA polymerase)

Promoter: a sequence of DNA which RNA polymerase attaches to begin transcription

Operator: a sequence that blocks action of RNA polymerase IF occupied by repressor protein

Structural Gene: DNA that codes for several related enzymes that direct production of product

The rest of the story….

In E. coli, the lac operon (controls breakdown of lactose) produces a repressor that binds to operator region, so RNA polymerase can’t transcribe genes that code for enzymes to breakdown and use Lactose.

But….when Lactose is present, binds with repressor, so RNA polymerase is able to transcribe proteins

So….is called an inducible enzyme, because the substance turns on the gene

And still more…

trp operon (enzymes for breakdown of tryptophan), produces inactive repressor that doesn’t bind to operator, so RNA polymerase proceeds. When tryptophan is available from environment, E. coli no longer has to make it, so tryptophan reacts' with the inactive repressor to make it active, acts as co repressor

Called repressible enzymes

Genome Organization at the DNA Level

Genome is plastic (changeable) in ways that affect availability of specific genes for expression

Some genes only work in certain cells, at certain time, in certain conditions (heterochromatin)

Changing Genome

Genome Organization

Repetitive-noncoding sequences account for much of genomeThink these introns protect DNA

from shortening during replication

Gene amplification increases selective DNA synthesis at certain time in development

Genome Organization

Some cancer cells have multiple copies which allows resistance to drugs and therapy

Rearrangement of GenomeTransposons move DNA within

genome to increase or decrease protein production

Genome Organization

ImmunoglobulinsDuring cellular differentiation,

rearrange the DNA that encodes antibodies, allows to recognize non self, become b-lymphocytes (white blood cells)

Mutation

Mistakes in genetic transmission

1 in 1x106 genes in meiosis and mitosis

Alteration in number and structure of chromosomes

Alteration in specific allele

Chromosomal Mutation

Nondisjuction: where sister chromatids fail to separateAnueploidy-abnormal number of a

certain chromosomeTrisomic, Monosomic

Polyploidy-two or more complete sets

Chromosomal Mutation con’t

Triploidy, Tetraploidy

Structure:Deletion, Duplication,

Translocation, Inversion

Gene Mutations

Mutations that effect a single gene or nucleotideFramshift, Gene Point

Cancer…not funny

Results from genetic changes that effect the cell cycle

Lack controls of growth and division in somatic cells

Caused by a mutation of a normal gene or by a viral agent

More Cancer….

Random and Spontaneous

Some Environmental causesVirususCarcinogons

Oncogene- cancer causing gene

And still more Cancer….

Whatever the cause, the mechanism is still the sameA mutation of the Genes that control growth and tumor suppressionNormally, more than one oncogene is mutated to cause cancer

Viruses and Cancer

Transform cells by inserting viral nucleic acids into host DNA

Is a permanent addition

15% of human cancers worldwide

Examples:

Viral Cancers

Retrovirus-Leukemia

Hespervirus-Mononuclous

Papvavorius-Cervical Cancer

Hepatitus B- Liver Cancer

DNA Technology

Practical goal is the improvement of human health and food production

Allows gene to be moved between species

Prodution of antibiotics, antibodies, fermentated products

Cloning

Recombinant DNA Technology

Technique used for recombining genes from different sources In Vitro and transferring this recombinant DNA into a cell to be expressed

Started in ’75 with Bacteria

Recombinant DNA con’t

Allows genes to move across species barriers

Allows constuction of recombinant DNA

Methods for purifying DNA and proteins

Recombinant DNA con’t

Vectors for carrying recom DNA into cells and replicating it

Techniques for determining nucleotide sequences of DNA

Restriction Enzymes

Major tools of recombination

Cut DNA into short segments at specific points

The the sticky ends (single stranded overlaps)of the DNA form bonds with the compliment

Restriction Enzymes

The recombinant DNA is carried by VectorsVectors are generally either bacterial

plasmids, or viruses

Then the DNA is incorporated

Polymerase Chain Reaction

Called PCRThis quickly amplifies DNA In VitroUsed in crime scenes, prenatel diagnosis from single cell, also with the wolly mammoth, the idea behind Jurassic Park

The Genome Project

Done by a variety of methods…..

Linkage mapping using markers

Physical mapping by cutting DNA into identifiable fragments, then overlaps

Sequencing DNA-by PCR, Chromosome Walking

Chromosome walking

Human\\Library\sys\HOME\t-mahan\Download video\Genome Timeline.exe Genome Timeline

The Human Genome Project

The Human Genome Project

Why Analyze The Genome?

Confirms Evolutionary Connection to Distant Organisms

To Study gene expression to determine which genes are active at certain species of development

Why Analyze The Genome?

To determine gene function to show mutation effects on protein productionHelps to understand metabolic

abnormalities

Applications of Genome Data

Genetic Counseling for prospective parentsShow possible traits by figuring

probability after studying possible recombinants

Carrier Recognition- of Parents

Application con’t- Counseling

Fetal TestingAmniocentisis- 14-16 wk,

looking for specific chemicals in amnionic fluid

PCR amplification for gene presence

Application con’t- Counseling

Fetal Testing con’tChronic Villus Sampling (CVS)-

down by karyotyping at 8 to 10wks

Applications- Gene Therapy

Replace or Supplement defective genes with functional

Normal genes introduced into Somatic cellsBut…..can we control protein

production?Does new harm other cells?

Applications- Gene Therapy

Pharmaceutical applicationsHuman InsulinGrowth HormoneTissue plasminogen activator (TPA)Engineer protein blocks to mimic or

block surface receptors (HIV)

Applications- Gene Therapy

Pharmaceutical applications con’tVaccines- Harmless variant or

derivative of a pathogen that stimulates the immune system to fight the pathogen

Two types- Inactive, ActiveRecombinant DNA techniques used to

produce

Applications- Forensics

DNA Fingerprinting- marker testing

PCR to amplify small samples

But…….what do we do with the DNA data gathered??????

How reliable?

Things that make you go….hmmmm

Should Genome Factor for jobs?

Who gets to examine your genes?

Costs? Insurance gets the bill?

Are vectors safe?

Who approves new products?

Who do we test them on?