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1: Genetic Scientists
November-15-13 5:06 PM

Friedrich Meischer Discovered that the nuclei of cells contain large quantities of a substance that does not act like protein He called the substance nuclein because it was found in the nucleus of the cell. Now the substance is called deoxyribonucleic acid.

Frederick Griffith Discovered that mice died after being injected with a mixture of heat killed S-strain bacteria and living R-strain bacteria. He concluded that the S-strain passed its deadly properties to the live non-pathogenic R-strain. Called it the transforming principle because the S-strain must have transformed something in the R-strain to make them deadly. Avery and his colleagues prepared identical extracts of the heat killed S-strain by growing the cells in liquid culture, isolating the bacteria, disrupting the cell membranes and collecting the cell content. One enzyme was added at a time. One enzyme was added to destroy protein, the second was added to destroy RNA and the third one to destroy DNA. Each enzyme-treated extract was mixed with the live R-strain cells. The only extract that did not cause any transformation in the R-strain was the third enzyme treated extract. The conclusion was that DNA was what caused the transformation. Hershey and Chase Experiment used bacteriophages, which are viruses that infect bacteria. They have an inner nucleic acid core and an outer protein core, called a capsid. Aimed to determine which part of the virus-the DNA or the protein-enters the bacterial cell and produces more viruses Since proteins contain sulphur but DNA does not, they introduced a radioactive source of sulfur into the protein of the virus. The DNA was labeled with phosphorus since, protein does not contain phosphorus Experiment One: A virus with radioactively labeled DNA was allowed to infect E.coli bacteria. The bacteria was then put in a blender to take the bacteriophage ghosts off the surface. The material was centrifuged to separate the infected bacterial cells (formed a pellet at the bottom) and the liquid medium (contains the bacteriophage ghost). The radioactive DNA was in the bacteria not in the liquid. Experiment Two: A virus with radioactively labeled protein was allowed to infect E.coli bacteria. The bacteria was then put in a blender to take the bacteriophage ghosts off the surface. The material was centrifuged to separate the infected bacterial cells (formed a pellet at the bottom) and the liquid medium (contains the bacteriophage ghost). The radioactive protein was in the liquid not in the bacteria. In conclusion was that viral DNA was transferred to the bacterial cells and that viral DNA held the genetic information needed for the viruses to reproduce.

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Phoebus Levene Isolated two types of nucleic acid: RNA (five carbon sugar) and DNA (five carbon sugar without one oxygen) Proposed that RNA and DNA are made up of different units called nucleotides. Each nucleotide would be composed of one of four nitrogen bases, a sugar molecule and a phosphate group.
There are four types of nitrogen-bases all linked together by covalent bonds. They are categorized into two different forms: purines and pyrimidines. The purine bases are adenine and guanine. They have two fused rings in there chemical structures. The pyrimidine bases are cytosine and thymine. They have a single ring in their chemical structures. In RNA it is uracil instead of thymine.

Erwin Chargaff Showed that there is variation in the composition of nucleotides among different species. Demonstrated that all DNA maintains certain properties, even though the composition varies. He observed that the nucleotides were present in characteristic proportions. Chargaff's rule; in DNA, the percent composition of adenine is the same as thymine, and the percent composition of cytosine is the same as guanine.
Linus Pauling Discovered that may proteins have a helix-shaped structure Rosalind Franklin Determined that DNA has a definite helix-shape. The structure has two regularly repeating patterns-one recurring at intervals of 0.34nm, and the other recurring at intervals of 3.4nm. Concluded that the nitrogenous bases were located on the inside of the helix structure, and the sugar-phosphate backbone was located on the outside, facing toward the watery nucleus of the cell

Watson and Crick Deduced that DNA has a twisted, ladder-like structure, called a double helix.

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Watson and Crick Deduced that DNA has a twisted, ladder-like structure, called a double helix.

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5.1: DNA Structure

November-16-13 1:29 PM

There are two polynucleotide strands twist around each other to form the double helix. The strands are comprised of alternating phosphate groups and sugars. The bases are attached to the sugar and protrude inwards. There is a constant total distance between backbones The purine molecule is always paired with the pyrimidine molecule. A-T and C-G. This is called complementary base pairing Hydrogen bonds link each complementary base pair. A and T share two and C and G share three. This makes C and G slightly stronger.

The two strands of DNA are antiparallel. At the end of a DNA molecule, the 5' end of one strand of DNA lies across from the 3' end of the complementary strand. The 5' and the 3' come from the numbering of the carbons on the deoxyribose sugar. The phosphate group is on the 5' carbon and the OH group is on the 3' carbon.

The genome is the complete genetic makeup of an organism; an organisms total DNA sequence The functional units of DNA are genes because their sequences code for the production of specific proteins or RNA For most prokaryotes genetic material is in the form of a circular, double stranded DNA molecule. Bacterial chromosome is tightly packed into the nucleoid. Specialized proteins that bind to the bacterial DNA help fold sections of the chromosome into loop-like structures DNA supercoiling is when the formation of additional coils in the structure of DNA due to twisting forces on the molecule. In bacteria, the amount of supercoiling is controlled by two enzymes: topoisomerase I and topoisomerase II. Antibacterial drugs stop the functionality of these enzymes. Some prokaryotes have one or two small, circular or linear DNA molecules called plasmids. They are not part of the nucleoid and carry non-essential genes. Most prokaryotes only carry one copy of their genes, so they are haploid organisms. Their genomes contain very little nonessential DNA.

Regulatory sequences are sections of DNA sequences that determine when certain genes and the associated cell functions are activated. For eukaryotic cells, the total amount of DNA is much greater than prokaryotic cells. DNA is also contained with the nucleus. Eukaryotes have to fold much more than prokaryotes. There are different levels of organization that takes place with each chromosome. A) DNA associates with proteins, histones, to form a repeated series of structures called nucleosomes. Each
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A) DNA associates with proteins, histones, to form a repeated series of structures called nucleosomes. Each nucleosome is composed of double-stranded DNA that is wrapped around a group of eight histone proteins. These nucleosome structures are connected by regions called linker DNA. B) Coiling of the nucleosomes with the help of H1 histone proteins, into which is often called a 30nm fibre. DNA is compacted 50 times C) Formation of radial loop domains of the 30nm fibre. They are anchored to a scaffold of proteins in the nucleus. Genetic material appears as a mass of long, intertwined strands known as chromatin. The 30nm fibre as looped domains, called euchromatin (can undergo further compacting) and heterochromatin. Most eukaryotes are diploid which means they contain two copies of each gene. However some may be haploid and can only carry one. Some may carry three or more copies of their genes. There is no correlation between an organisms complexity and genome size or number of proteincoding genes

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5.2 DNA Replication

November-16-13 6:53 PM

DNA replication is the process of producing two identical DNA molecules from an original, parent DNA molecule. Due to complementary base pairing each strand can serve as a template for the production for a new complementary strand There three main models for replication of DNA: Conservative model (two new daughter strand from the parent templates, with two new strands joining to create a new double helix) Semi-conservative model (each new DNA molecule would contain one strand of the new complementary DNA molecule and one new parent strand. Each new DNA molecule would conserve half of the strand of the original molecule) The dispersive model (proposed that the parental DNA fragments were broken into fragments and that both strands of DNA in each of the daughter molecules were made up of an assortment of parental and new DNA)

Matthew Meselson and Franklin Stahl reasoned that the proposed models for DNA replication could be tested if they could distinguish between the original parent strand and the new daughter strands. They used two different isotopes of nitrogen to label the DNA in the cell: the common form of nitrogen, 14N, and a rarer form of heavy nitrogen, 15N. The light and heavy forms of nitrogen were used because it was easy to separate the different DNA strands based on how much of each isotope is present in a newly synthesized DNA molecule. Bacterial cells were grown for several generation in medium containing a heavy isotope of nitrogen. Therefore they contained heavy nitrogen. Cell were transferred to a new medium containing normal, light nitrogen. At various times after, samples were collected. The DNA was extracted from the cells and put in a centrifuge so that a concentration gradient could be observed. Heavier sediments form rings toward the bottom. After one generation of growth in 14N medium, the bacteria yielded a single band of DNA with a density between that of 14N-DNA and 15N-DNA indicating that one strand of each contained 15N After two generations of growth in 14N medium, the bacteria yielded two bands. One contained no 15N and the other contained 14N and 15N. There are three basic phases in replication that rely on

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There are three basic phases in replication that rely on the structural feature of DNA and a number of specialized proteins. 1) In the initiation phase, a portion of the DNA helix is unwound to expose the bases for new base pairing. 2) In the elongation phase, two new strands of DNA are assembled using the parent DNA as a template. The new DNA molecules - each composed of one strand of parent DNA and one strand of daughter DNA - re-form into double helices. 3) In the termination phase, the replication process is completed and the two new DNA molecules separate from each other. All these events take place simultaneously. Replication starts at a specific nucleotide sequence called the origin of replication. Several proteins bind to the DNA and unwind the double helix. The helicase enzyme cleave the hydrogen bonds that link the complementary base pairs together. Single -strand-biding proteins help to stabilize the newly unwound strands (they have a tendency to get back together). The topoisomerase II enzyme helps relieve the strain on the double helix sections ahead of the replication forks. Initiation creates an unwound, oval shape called the replication bubble, with two Y-shaped regions at each unwound area. The Y-area is called the replication fork. DNA polymerase III is the enzyme that catalyzes the addition of new nucleotides, to create a new strand of DNA that is complementary to a parental strand. The enzyme only attaches new nucleotides to the 3' OH end of a pre -existing chain of nucleotides. It can only synthesis a new strand from a parent strand in the 5' to 3' direction. When double-stranded DNA is separated the strands do not have free 3'hydroxyl end for DNA polymerase to begin at. Therefore, new DNA requires both strands to be started with short fragments of nucleotide sequences, complementary to the templates. For once strand, the leading strand, this only need to happen once. DNA polymerase will keep adding new nucleotides to the 3' end as it moves in the same direction as the replication fork. Synthesis of the other end, lagging strand, requires the enzyme to move in the opposite direction. This needs to occur in sho rt fragments and in a discontinuous manner. These short segments are called the Okazaki fragments. RNA primers are needed to start the replication. RNA primers are synthesized by an enzyme called primase. DNA polymerase extends the strand by adding new nucleotides to the 3'end of the primer. Once each primer is in place, a new DNA fragment is generated at the end of each prmer. The result is short segments containing uracil instead of thymine. Another enzyme, DNA primase I, is needed to replace the uracil. Then the two strands are joined together by DNA ligase.

As soon as the newly formed strands are complete, they rewind automatically. The DNA complex at each replication fork that carries out replication is referred to as the replication machine. The termination phase occurs upon the new DNA strands completion and the two new DNA molecules separate from each other. One type of error that can take place is a mismatching between the new nucleotide and the template. Another type of problem that can occur is due to strand slippage, which causes either additions or omissions of nucleotides. This type of error can result from either the newly synthesizing strand looping out, allowing addition of an extra nucleotide or the looping out of the template strand, resulting in a nucleotide not being added where it should. Enzymes DNA polymerase I and II proofread each nucleotide and recognize whether or not the correct nucleotide has been added. If it detects an error it will halt production and fix it.
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has been added. If it detects an error it will halt production and fix it. Another mechanism for correcting errors is called mismatch repair. Mismatching pairs cause deformities in the molecule. Special enzymes bind to these parts of the daughter cell and repair them. Telomere is a repetitive section of DNA, near the end of each chromosome; this helps to protect from loss of information during replication of the linear DNA in eukaryotic cells

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