BIS 2A Ch. 13
How much primer is needed for the synthesis of the leading strand? Okazaki fragments?
-1 -one per fragment
3 DNA repair mechanisms
1. A proofreading mechanism corrects errors in replication as DNA polymerase makes them. 2. A mismatch repair mechanism scans DNA immediately after it has been replicated and corrects any base-pairing mismatches. 3. An excision repair mechanism removes abnormal bases that have formed because of chemical damage and replaces them with functional bases.
what are the 4 key structures of DNA?
1. Double stranded helix with a sugar-phosphate backbone on the outside and base pairs lined up on the inside. 2. DNA is usually a right-handed helix. If you curl the fingers of your right hand and point your thumb upward, the curve of the helix follows the direction of your fingers, and it winds upward in the direction of your thumb. 3. DNA is antiparallel 4. DNA has major and minor grooves in which the outer edges of the nitrogenous bases are exposed.
What are the 2 steps in DNA replication?
1. The DNA double helix is unwound to separate the two template strands and make them available for new base pairing. 2. As new nucleotides form complementary base pairs with template DNA, they are covalently linked together by phosphodiester bonds, forming a polymer whose base sequence is complementary to the bases in the template strand.
What are the steps in PCR?
1. The first step involves heating the reaction mixture to near boiling point, to separate (denature) the two strands of the DNA template. 2. The reaction is then cooled to allow the primers to bind (or anneal) to the template strands. 3. Next, the reaction is warmed to an optimum temperature for the DNA polymerase to catalyze the production of the complementary new strands.
4 functions of DNA based on the double helix structure
1. The genetic material stores an organism's genetic information. With its millions of nucleotides, the base sequence of a DNA molecule can encode and store an enormous amount of information. Variations in DNA sequences can account for species and individual differences. DNA fits this role nicely. 2. The genetic material is susceptible to mutations in the information it encodes. For DNA, mutations might be simple changes in the linear sequence of base pairs. 3. The genetic material is precisely replicated in the cell division cycle. Replication could be accomplished by complementary base pairing, A with T and G with C. 4. The genetic material (the coded information in DNA) is expressed as the phenotype. This function is not obvious in the structure of DNA. However, the nucleotide sequence of DNA is copied into RNA, which uses the coded information to specify a linear sequence of amino acids—a protein. The folded forms of proteins determine many of the phenotypes of an organism.
What were the 2 ways Avery identified the substance causing bacterial transformation?
1.Eliminating other possibilities. Cell-free extracts containing the transforming substance were treated with enzymes that destroyed candidates for the genetic material, such as proteins, RNA, and DNA. When the treated samples were tested, the ones treated with RNase and protease (which destroy RNA and proteins, respectively) were still able to transform R-type bacteria into the S-type. But the transforming activity was lost in the extract treated with DNase (which destroys DNA) 2. Positive experiment. They isolated virtually pure DNA from a cell-free extract containing the transforming substance. The DNA alone caused bacterial transformation. 2.
Templates
A molecule or surface on which another molecule is synthesized in complementary fashion, as in the replication of DNA.
Replication forks
A point at which a DNA molecule is replicating. The fork forms by the unwinding of the parent molecule.
Polymerase chain reaction (PCR)
An enzymatic technique for the rapid production of millions of copies of a particular stretch of DNA where only a small amount of the parent molecule is available.
Telomerase
An enzyme that catalyzes the addition of telomeric sequences lost from chromosomes during DNA replication.
Primase
An enzyme that catalyzes the synthesis of a primer for DNA replication.
DNA polymerase
Any of a group of enzymes that catalyze the formation of DNA strands from a DNA template.
Complementary base pairing
Base pairing with one purine and one pyrimidine
How does DNA polymerase work so fast?
DNA polymerases are processive- catalyze the formation of many Phosphodiester linkages each time they bind to a DNA molecule
Origin of replication
DNA sequence at which helicase unwinds the DNA double helix and DNA polymerase binds to initiate DNA replication
DNA ligase
Enzyme that unites broken DNA strands during replication and recombination.
Explain Griffith experiment with smooth and rough strain of bacteria experiment
Griffith was working with two strains of pneumococcus: Cells of the S strain produced colonies that looked smooth (S). Covered by a polysaccharide capsule, these cells were protected from attack by a host's immune system. When S cells were injected into mice, they reproduced and caused pneumonia (the strain was virulent). Cells of the R strain produced colonies that looked rough (R), lacked the protective capsule, and were not virulent. When Griffith inoculated mice with heat-killed S-type pneumococcus cells, the cells did not produce infection. However, when he inoculated other mice with a mixture of living R-type cells and heat-killed S-type cells, to his astonishment, the mice died of pneumonia (Figure 13.1). When he examined blood from these mice, he found it full of living bacteria—many of them with characteristics of the virulent S strain! Griffith concluded that in the presence of the dead S-type pneumococcus cells, some of the living R-type cells had been transformed into virulent S cells. These cells were able to grow in the bodies of the mice, causing pneumonia and multiplying in the blood. The fact that these S-type cells reproduced to make more S-type cells showed that the change from R-type to S-type was genetic.
Meselson and Stahl prove DNA replication is semi conservative experiment
Heavy nitrogen (15N) is a rare, nonradioactive isotope that makes molecules containing it denser than chemically identical molecules containing the common isotope 14N. Two cultures of the bacterium E. coli were grown for many generations, one in a medium containing 15N and the other in a medium containing 14N. When DNA extracts from the two cultures were combined and centrifuged in a solution of cesium chloride, which forms a density gradient under centrifugation, two separate bands of DNA formed in the centrifugation tube. The DNA from the 15N culture was heavier than the DNA from the 14N culture, so it formed a band at a different position in the density gradient. Next, Meselson and Stahl grew another E. coli culture in 15N medium, then transferred the bacteria to normal 14N medium and allowed them to continue growing. The cells replicated their DNA and divided every 20 minutes. Meselson and Stahl collected some of the bacteria at time intervals and extracted DNA from the samples. The results can be explained only by the semiconservative model of DNA replication.
Single strand binding protein
In DNA replication, a protein that binds to single strands of DNA after they have been separated from each other, keeping the two strands separate for replication.
Leading strand
In DNA replication, the daughter strand that is synthesized continuously.
Lagging strand
In DNA replication, the daughter strand that is synthesized in discontinuous stretches.
Genetic marker
In general, a DNA sequence such as a single nucleotide polymorphism whose presence is correlated with the presence of other linked genes on that chromosome.
Transfection
Insertion of recombinant DNA into animal cells.
Okazaki fragments
Newly formed DNA making up the lagging strand in DNA replication. DNA ligase links Okazaki fragments together to give a continuous strand.
Where are nucleotides added to in DNA replication?
On the growing new strand at the 3' end, end with free hydroxyl
Antiparallel
Pertaining to molecular orientation in which a molecule or parts of a molecule have opposing directions.
sliding DNA clamp
Protein complex that keeps DNA polymerase bound to DNA during replication.
How can DNA become damaged?
Radiation or chemicals
Telomeres
Repeated DNA sequences at the ends of eukaryotic chromosomes.
Primer
Strand of nucleic acid, usually RNA, that is the necessary starting material for the synthesis of a new DNA strand, which is synthesized from the 3′ end of the primer
Hershey and Chase experiment with bacteriophages T2
T2 phage consists of a DNA core packed inside a protein coat. When it attacks a bacterium, part of the virus enters the bacterial cell. About 20 minutes later, the cell bursts, releasing dozens of particles that are virtually identical to the infecting virus particle. Clearly the virus is somehow able to convert the host cell from its own genetic program into a viral replication machine. Hershey and Chase set out to determine what part of the virus—DNA or protein—enters the host cell to bring about this genetic change. To trace the two components of the virus over its life cycle, the scientists labeled each component with a specific radioisotope: Proteins were labeled with radioactive sulfur. Proteins contain some sulfur, but DNA does not. Sulfur has a radioactive isotope, 35S. Hershey and Chase grew bacteriophage T2 in a bacterial culture in the presence of 35S, so the proteins of the resulting viruses contained the radioisotope. DNA was labeled with radioactive phosphorus. DNA contains a lot of phosphorus, whereas proteins contain little or none. Phosphorus also has a radioisotope, 32P. The researchers grew another batch of T2 in a bacterial culture in the presence of 32P, thus labeling the viral DNA with 32P. Hershey and Chase used these radioactively labeled viruses in their experiments. In one experiment, they allowed 32P-labeled bacteriophage to infect bacteria; in the other, the bacteria were infected with 35S-labeled bacteriophage. After a few minutes they agitated each mixture vigorously in a kitchen blender, stripping away the parts of the viruses that had not penetrated the bacteria, without bursting the bacteria. Then they separated the bacteria from the rest of the material (remains of viruses) in a centrifuge. The result was that the bacterial cells in the centrifuge pellet contained most of the 32P, and the supernatant fluid with the viral remains contained most of the 35S. These results indicated that it was the DNA that had been transferred into the bacteria, and that DNA was the molecule responsible for redirecting the genetic program of the bacterial cell.
what is Chargaff's discovery?
That the amount of adenine equals the amount of thymine and the amount of cytosine equals the amount of guanine. This provided an important clue about the way the bases were arranged in a DNA double helix
What are crucial observations about semi conservative replication evidence?
The crucial observations demonstrating this model were that all the DNA at the end of the first generation was of intermediate density, while at the end of the second generation there were two discrete bands: one of intermediate and one of light DNA. If the conservative model had been true, there would have been no intermediate density DNA. If the dispersive model were correct, then the DNA would all have been intermediate for the first few generations, with the single intermediate band becoming progressively lighter.
3' end
The end of a DNA or RNA strand that has a free hydroxyl group at the 3′ carbon of the sugar (deoxyribose or ribose).
How do the 4 possible configurations of the h bonded base pairs in the major and minor grooves show that they can interact with other molecules?
The exposed outer edges of the base pairs are accessible for additional hydrogen bonding. Note that the arrangements of unpaired atoms and groups differ in A-T and G-C pairs. Thus, the surfaces of the A-T and C-G base pairs are chemically distinct, allowing other molecules such as proteins to recognize specific base pair sequences and bind to them. The binding of proteins to specific base pair sequences is the key to protein-DNA interactions, which are necessary for the replication and expression of the genetic information in DNA.
Semiconservative replication
The way in which DNA is synthesized. Each of the two partner strands in a double helix acts as a template for a new partner strand. Hence, after replication, each double helix consists of one old and one new strand.
Why do many cell lineages not last the entire lifetime of the organism?
Their telomeres are lost
5' end
end of a DNA or RNA strand that has a free phosphate group at the 5′ carbon of the sugar (deoxyribose or ribose).
DNA helicase
enzyme that unwinds the double helix
Dispersive replication
replication that would produce 2 molecules with old and new DNA interspersed along each strand
Conservative replication
replication where the original double helix serves as a template for, but does not contribute to, a new double helix