DNADNAThe Code of LifeThe Code of Life

The Molecular Basis of

Inheritance

DNA Deoxyribonucleic acidThe information necessary to sustain and perpetuate life is found within a molecule. This is the genetic material that is passed from one generation to the next---a blue print for building living organisms.

HistoryHistoryAlthough we now accept the idea that DNA is responsible for our biological structure,But in the early 1800s it was unthinkablefor the leading scientists and Philosophers that a chemical molecule could hold enough information to build a human. They believed that plants and animals had been specifically designed by a creator.

HistoryHistory

Charles Darwin is famous for challenging this view. In 1859 he published 'The Origin of Species‘ expressing that living things might appear to be designed, but were actually the result of natural selection.Darwin showed that living creatures evolve over several generations through a series of small changes.

HistoryHistory

In the 1860s Darwin's ideas were supported when genetics was discovered byGregor Mendel. He found that genesdetermine the characteristics a living thing will take. The genes are passed on to later generations, with a child taking genes from both its parents. The great mystery was where and how is this information stored?

HistoryHistory

The main conclusions made by Mandle were:*SEGREGATION:Inherited traits are controlled by genes, which are in pairs. When sex cells are created one gene from each pair goes into the gamete. When two gametes fuse at fertilization, the offspring has two copies of each gene—one from each parent.*INDEPENDENT ASSORTMENT:The genes for different traits are sorted into gametes independentlyof other genes. So one inherited trait is not dependent on another.*DOMINANCE:Where there are two different forms of a gene are present in a pea plant, the one which is dominant is the one that is observed.

HistoryHistorySearch for genetic material:In 1870, a German scientist named Friedrich Miescher had isolated the chemicalsfound in the nucleus. These were proteins and nucleic acids. While he found these nucleic acidsinteresting, and spent a great deal of time studying their chemical composition, he wasn’t alone inbelieving that proteins were more likely to be the chemicals involved in inheritance, because of theirimmense variability. They were made up of 20 different building blocks (amino acids), as opposed tothe mere 4 building blocks of nucleic acids.

HistoryHistorySearch for genetic material:In the early 1900s, Phoebus Levene, who also believed that proteins must be the chemicals ofinheritance, studied the composition of nucleic acids. He discovered that DNA is a chain ofnucleotides, with each nucleotide consisting of a deoxyribose sugar, a phosphate group and anitrogenous base, of which there were four different types. He proposed that the four different types ofnucleotide were repeated over and over in a specific order. This would make DNA a relatively simplerepeat sequence – no wonder DNA wasn’t considered to be smart enough to code for all of life!

HistoryHistorySearch for genetic material:

1928 Frederick Griffith: transforming principle

Search for genetic material:

HistoryHistory

It wasn’t until 1944 that Oswald Avery and his colleagues, who were studying the bacteria whichcauses pnuemonia, Pneumococcus, discovered by process of elimination that bacteria contain nucleicacids, and that DNA is the chemical which carries genes.Despite the conclusive results of Avery’s experiments, the theory of nucleic acids being the geneticmaterial was still not a popular one, but experimentsPerformed with viruses also showed that nucleicacids were the genetic material and this confirmed Avery’s work.

Search for genetic material:1952 - Hershey-Chase

Experiment

HistoryHistory

HistoryHistorySearch for genetic material:

Classic experiments for evidence Griffith: transformationHershey-Chase: DNA necessary

to produce more virus

Other supporting evidenceDNA volume doubles before cells

divideChargaff: ratio of nucleotides

A = T and G = C

The DiscoveryThe Discovery

The DNA molecule was discovered in 1951 by Francis Crick, James Watson and Maurice Wilkins using X-ray Diffraction. In Spring 1953, Francis Crick and James Watson, two scientists working atthe Cavendish Laboratory in Cambridge, discovered the structure of the DNA a double helix,or inter-locking pair of spirals, joined by pairs of molecules.

The DiscoveryThe Discovery

The seed that generated this was Watson’spresence at a conference in Naples in 1951, where an x-ray diffraction picture from DNA was shown by Maurice Wilkins from King’s College in London.This made a strong impression on Watson – the first indicationthat genes might have a regular structure.

HistoryHistorySearch for genetic material:

James Watson joined the unit (its first biologist) and began bytrying to crystallize myoglobin for Kendrew. The unsuccess of this left much time for discussion with Crick, whose office he was sharing, and the topic of DNA structure naturally arose –particularly how to determine it. They were inclined to follow the method of Pauling who had deduced the a-helical structure by building a model consistent with the x-ray patterns from fibrousproteins. Like proteins, DNA was built from similar units – the bases adenine (A) thymine (T) guanine (G) and cytosine (C),and so it seemed likely that DNA too had a helical structure. The publishedx-ray patterns of DNA were not very clear, and so contact was made with King’s.Watson attended a DNA colloquium there in November 1951, at which Rosalind Franklin described her results.

HistoryHistory

Search for genetic material:

Watson brought back a less-than-accurate accountto Cambridge, but with Crick produced a three-strandmodel structure only a week later. Invited to view this,Franklin pointed out that it was inconsistent with her results – it had thephosphate groups on the insidewhereas her results showed they were on the outside,and the water content was too low. The work at Cambridge stopped abruptly for a bit.

HistoryHistorySearch for genetic material:

In July 1952, Erwin Chargaff visited the unit and told of his 1947 findings that the ratios of A/T and G/C were unity for a wide variety of DNAs. Crick became convinced that base pairing was the key to the structure. Prompted by receiving a flawed manuscript on DNA structure from Pauling, Watson again visited King’s and Wilkins showed him a DNA x-ray pattern taken by Franklin of the pure B-form showing clear helical characteristics, plus the intense 10th layer line at 3.4A and a 20A equatorial reflection indicating the moleculardiameter. Perutz also showed them a report on the work of the King’s group which gave the space group of the crystalline A-form as C2, from which Crick deduced that there were two chains running in opposite directions.

HistoryHistorySearch for genetic material:

Watson began pursuing the idea of hydrogen bonding using cardboard cutouts of the four bases. He found that (A+T) and (G+C) could be bonded together to form pairs with very similar shapes. On this basis a model was built consistent with the symmetry and with Chargaff’s results, and a paper waspublished in April 1953 in Nature accompanied by ones from the Wilkins and Franklin groups at King’s. Watson and Crick’spaper ends with the oft-quoted line “It has not escaped ournotice that the specific pairing we have postulated immediatelysuggests a possible copying mechanism for thegenetic material”.

The EvidenceThe EvidenceSearch for genetic material:

James Watson and Francis Crick used this photo with other evidence to describe the structure of DNA.

X-ray diffraction photo of DNAImage produced by Rosalind Franklin

Watson and Crick with their

DNA model

The ScientistsThe Scientists

Francis Crick was born in 1916. He went toLondon University and trained as a physicist. After the war he changed the direction of his research to molecular biology. James Watson was an American, born in 1928, so aged only 24 when the discovery was made. He went to Chicago University aged only 15 and had already worked on DNA.

The Nobel PrizeThe Nobel Prize

Crick, Watson and Wilkins won the Nobel Prize for medicine in 1962. Maurice Wilkins was at King's College, London and was an expert in X-ray photography. His colleague, Rosalind Franklin, did brilliant work developing the technique to photograph a singlestrand of DNA. She received little recognition for thisat the time and died tragically of cancer in 1958, so could not be recognised in theNobel Award.

Watson & CrickWatson & Crick

What they deduced from:Franklin’s X-ray data• Double helix• Uniform width of 2 nm• Bases stacked 0.34 nm apart

Chargoff’s “rules”• Adenine pairs with thymine• Cytosine pairs with guanine

Watson & CrickWatson & Crick

What they came up with ontheir own:• Bases face inward, phosphates and sugars outward• Hydrogen bonding• Hinted at semi-conservative model for replication

KEY PLAYERS

Oswald Avery (1877-1955)

Microbiologist Avery led theteam that showed that DNA is the unitof Inheritance. One Nobel laureate has called the discovery "the historical platform of modern DNA research", and his work inspired Watson andCrick to seek DNA's structure.

KEY PLAYERS

Erwin Chargaff (1905-2002)

Chargaff discovered the pairing rules of DNA letters, noticing that A Matches to T and C to G. He laterCriticized molecular biology, the discipline he helped invent, as "the practice of biochemistry without a licence",and once described Francis Crick as looking like "a faded racing tout".

KEY PLAYERS

Francis Crick (1916- )

Crick trained and worked as a physicist, but switched to biology after the Second World War. After co-discovering the structure of DNA, he went on to crack the genetic code that translates DNA into protein. He now studies consciousness at California's Salk Institute.

KEY PLAYERS

Rosalind Franklin (1920-58)

Franklin, trained as a chemist, was expert in deducing the structure of molecules by firing X-rays through them. Her images of DNA - disclosed without her knowledge - put Watson and Crick on the track towards the right structure. She went on to do pioneering work on the structures of viruses. .

KEY PLAYERS

Linus Pauling (1901-94)

The titan of twentieth-century chemistry. Pauling led the way in working out the structure of big biological molecules, and Watson and Crick saw him as their main competitor. In early 1953, working without the benefit of X-ray pictures, he published a paper suggesting that DNA was a triple helix.

KEY PLAYERS

James Watson (1928- )

Watson went to university in Chicago aged 15, and teamed up with Crick in Cambridge in late 1951. After solving the double helix, he went on to work on viruses and RNA, another genetic information carrier. He also helped launch the human genome project, and is president of Cold Spring Harbor Laboratory in New York.

KEY PLAYERS

Maurice Wilkins (1916- )

Like Crick, New Zealand-born Wilkins trained as a physicist, and was involved with the Manhattan project to build the nuclear bomb. Wilkins worked on X-ray crystallography of DNA with Franklin at King's College London, although their relationship was strained. He helped to verify Watson and Crick's model, and shared the 1962 Nobel with them.

StructureStructure

StructureStructure

StructureStructure

StructureStructure

StructureStructure

StructureStructure There are 4 different nucleotides in DNA

AdenineAdenine pairs with ThymineThymineGuanineGuanine pairs with CytosineCytosine

StructureStructure

AdenineAdenine pairs with ThymineThymineGuanineGuanine pairs with CytosineCytosine

StructureStructureDoes DNA fit the requirements of a hereditary material?

Requirement DNA componentHas biologically useful information to make protein

Genetic code: 3 bases code for 1 amino acid (protein)

Must reproduce faithfully and transmit to offspring

Complementary bases are faithful; found in germ cells

Must be stable within a living organism

Backbone is strong covalent bonds; hydrogen bonds

Must be capable of incorporating stable changes

Bases can change through known mechanisms

Protein SynthesisProtein Synthesis

DNA carries the instructions for the production ofproteins.A protein is composed of smaller molecules called amino acids, and the structure and function ofthe protein is determined by the sequence of its amino acids. The sequence of amino acids, in turn, is determined by the sequence of nucleotide bases in the DNA. A sequence of three nucleotide bases, called a triplet, is the genetic code word, or codon, that specifies a particular amino acid.

Protein SynthesisProtein Synthesis

Protein synthesis begins with the separation of a DNA molecule into two strands. In a process called transcription, a section of the sense strand acts as a template, or pattern, to produce a new strand called messenger RNA (RNA). The RNA leaves the cell nucleus and attaches to the ribosomes, specializedcellular structures that are the sites of protein synthesis.Amino acids are carried to the ribosomes by another type of RNA, called transfer (RNA). In a process called translation, the amino acids are linked together in a particular sequence, dictated by the RNA, to form a protein.

ReplicationReplication

Before replication, the parent DNA molecule has 2 complementary strands

First the 2 strands separate

Each “old” strand serves as a template to determine the order of the nucleotides in the new strand

Nucleotides are connected to form the backbone; now have 2 identical DNA molecules.

ReplicationReplication

Helicase unwinds the molecule Single-strand binding protein stabilized ssDNA Primase initiates the replication with RNA DNA polymerase extends the new DNA Second DNA polymerase removes the RNA DNA ligase joins all the fragments

DNA Replication is simple, but it takes a large team of enzymes and proteins to carry out the process:

1971-Smith & 1971-Smith & NathansNathans

Discovery of restriction endonucleasesHamilton Smith• Discovered HindII inHaemophilus influenzaeDaniel Nathans• Used HindII to make firstrestriction map of SV40

1972- Paul BergProduces first recombinant DNA using EcoRI 1973 -Boyer, Cohen & ChangTransform E. coli with Recombinant plasmid

1977- Genentech, Inc.• Company founded byHerbert Boyer and RobertSwanson in 1976• Considered the advent ofthe Age of BiotechnologyFirst human protein (somatostatin) producedfrom a transgenic bacterium.• Walter Gilbert and Allan Maxam devise a method for sequencing DNA.

1978• David Botstein discovers RFLP analysis

1980• U.S. Supreme Court rules that life forms can be patented• Kary Mullis develops PCR. Sells patent for $300M in 1991

1981• First transgenic mice produced

1982• The USFDA approves sale of genetically engineered human insulin

1983• An automated DNA sequencer is developed• A screening test for Huntington’s disease is developed usingrestriction fragment length markers.

1984• Alec Jeffreys introduces technique for DNA fingerprintingto identify individuals

1985• Genetically engineered plants resistant to insects, viruses,and bacteria are field tested for the first time• The NIH approves guidelines for performing experiments ingene therapy on humans

1987

• invention of YACs (yeast artificial chromosomes) asexpression vectors for large proteins

1989• National Center for Human Genome Research created tomap and sequence all human DNA by 2005.

1990• UCSF and Stanford issued their 100th recombinant DNApatent and earning $40 million from the licenses by 1991.• BRCA-1 discovered• First gene therapy attempted on a four-year-old girl with an inherited immune deficiency disorder.

1992• U.S. Army begins "genetic dog tag" program

1994• The Flavr Savr tomato gains FDA approval• The first linkage map of the human genome appears

1995• The first full gene sequence of a living organism iscompleted for Hemophilus influenzae.• O.J. Simpson found not guilty despite DNA evidence

1996

• The yeast genome, containing approximately 6,000 genes and fourteen million nucleotides, is sequenced.

1997• Dolly cloned from the cell of an adult ewe• DNA microarray technology developed

1997

•The genome of the bacterium E. coli, a classic model organism for studying microbiological and molecular genetic mechanisms, and a natural symbiont in the human digestive tract, is completely sequenced, revealing about 4,600 genes among about four and one-half million nucleotides.

1998

The genome of a nematode worm Caenorhabditis elegans, a key model organism for investigating genetic regulation of development, is sequenced, revealing approximately 18,000 genes among some 100 million nucleotides of DNA sequence.

1999

• 1,274 biotechnology companies in the United States• At least 300 biotechnology drug products and vaccinescurrently in human clinical trials• Human Genome Project is on time and under budget, the complete human genome map expected in five years or less

1999

•Jesse Gelsinger, an eighteen year-old with a genetic disorder affecting liver metabolism, dies from an immune reaction to a gene therapy treatment. This tragic event slows gene therapy applications and results in greater scrutiny and caution toward the growing number of gene therapy research trials.

1999

•The first complete sequence of a human chromosome (number 22) is completed by the public genome project and is published. This step indicates that the genome project is proceeding ahead of schedule, and also shows a surprisingly small number of genes (about 300) relative to the anticipated 100,000 or so for all twenty-four human chromosomes (twenty-two chromosomes called autosomes shared equally by males and females, plus the X-chromosome which is paired in females but occurs in a single copy in males, plus the Y-chromosome that is unique to males).

2000

• Celera sequences the genome of the fruitfly (Drosophila melanogaster), identifying approximately 13,000 genes among 170 million nucleotides.•First plant genome sequenced (Arabidopsis thaliana) from the mustard family. The Arabidopsis genome consists of about 100 million nucleotides, and approximately 20,000 genes, indicating that at the molecular genetic level, plant and animal genomes are about equally complex.

2000

•"Golden rice," a genetically engineered strain of rice manufactures its own vitamin A. Golden rice is created by Ingo Potrykus, plant geneticist, and his colleagues to help alleviate severe health problems in many areas of the world caused by vitamin A deficiency.

2001

•In mid-February, the journal SCIENCE publishes an analysis of the Celera Human Genome Project, and the journal NATURE publishes an analysis of the public Human Genome Project. Both revealed a surprisingly small number of human genes, estimated jointly at about 30,000 to 35,000, barely more than a worm, fruitfly, or plant. Both show that only about 2 percent of our DNA actually codes for amino acid sequences of proteins, and both identify many sequences of unknown function and variable length present in multiple copies making up approximately half the genome.

ExtractionExtractionEach human cell has enough DNA to code for all the traits in the human body. If the DNA in one cell was stretched out, how long would it be? Do the math!

There are 6 X 109 base pairs/cellEach base pair is 0.34 X 10-9 meters long

A human body has approximately 75 trillion cells. If the distance to the sun is 150 X 109 meters, how many round trips could your DNA make?

Answer: 2 meters

Answer: 500 trips

ExtractionExtraction

DNA from kiwi fruit