Molecular Basis of Inheritance Flashcards
After Morgan identified that genes are located on chromosomes, what was the next step for researchers to figure out in regards to the genetic material?
Which part of the chromosome were the genetic material: DNA or protein.
What was very important in determining the genetic material of chromosomes? What organism was used to discover this?
Choosing the right experimental organism.
Studying bacteria and viruses that kill them (bacteriophage)
Who began the research that led to the discovery of DNAs genetic role?
Frederick Griffith in 1928
What organism did Griffith work with in his research? What were the two strains? What was the difference between the two strains?
Streptococcus pneumoniae
S strain is pathogenic
R strain is harmless
What were the results of Griffiths experiments with stretococcus pneumoniae? What did he call the phenomenon that occured, could he tell if the DNA or protein was causing it?
How could he tell whether or not the bacteria was pathogenic?
He heat-killed the pathogenic S strain of bacteria and placed the remains with living R strain bacteria
This resulted in some of the living R cells to become pathogenic
This phenomenon he called transformation, which was eventually was found to be a change in the genotype and phenotype due to the assimilation of external DNA. At the time he still didn’t know whether the DNA or proteins caused the transformation.
He would place his bacteria in mice and see if they killed the mouse or if it remained healthy. S cells killed the mouse, R cells did not. Heat-killed S cells did not kill the mouse. Heat-killed S cells mixed with living R cells killed the mouse.
R CELLS TRANSFORMED INTO S CELLS
How did, in 1944, Oswald Avery, Maclyn McCarty, and Colin MacLeod come up with the idea that DNA had to do with the transformation phenomenon that changes harmless bacteria into pathogenic bacteria?
The used enzymes that broke down different parts of the chromosome, either DNA or proteins, as well as RNA. Like DNAase, proteases, and RNAase.
After one of these were administered they saw whether or not transofrmation still occured with the remaining products.
It was found that only when DNA remained did transformation still occur.
The scientists initially using these in molecular genetic research did not know this, but how do bacteriophages work?
They insert their DNA into the bacteria, essentially they infect them.
How did, in 1952, Alfred Hershey and Martha Chase show that DNA is the genetic material? What did they use?
They used a phage (bacteriophage) called a T2 and determined what its genetic material is.
They did this by finding whether the DNA or the protein is what entered the host cell during the course of its infection. Whatever entered was its genetic material.
What were the specifics of Alfred Hershey and Martha Chase experiments with T2 phage to determine its genetic material? Basically describe the experiment step by step.
- They labeled either the protein or the DNA of the T2 with a radioactive isotope (sulfur for protein and phosphorus for DNA) and separated them into two batches.
- Each batch of T2 was introduced to nonradioactive E. coli cells to allow them to be infected by each batch separately.
- After allowing time for infection, the mixture was agitated in a blender to separate the remaining phage parts (soupinate) from the bacteria (pellet).
- The pellet at the bottom of the mixture was the bacteria and the remaining phage parts suspended above it in the liquid soupinate.
- The radioactivity of the pellet and the liquid (soupinate) above for each batch was measured.
- It was found that the DNA labeled batch had radioactivity in the pellet while the protein labeled bactch had radioactivity in only the liquid.
- This means that the DNA was the material that entered the cell, not the protein, and therefore the DNA must have reprogrammed it, making a strong argument that the DNA contained the genetic material.
- It was also shown that after the infection had run its course the bacteria released new phages that contained radioactive DNA, showing its ongoing role in the infection process.
Why did different radioactive isotopes have to be used for Hershey and Chase experiment?
Each isotope was an element specific to each molecule being tested.
How did Erwin Chargaff, in the 1950s, make the argument even stronger that DNA is the genetic material?
He was able to report that DNA composition varies species to species.
What are the two components of Chargaffs rules?
- DNA composition varies between species
- A = T and G = C in regards to nitrogenous bases.
A DNA polymer consists of? Was this known around Chargaffs time?
Pentose sugar (deoxyribose)
Phosphate group
Nitrogenous base
This was known in the 50s.
Why are Maurice Wilkins and Rosalind Franklin important in the scientific community?
They produced a picture of a DNA molecule using X-ray crystallography.
How was the image of DNA produced by X-ray crystallography? What was the significance of this image and Franklins conclusions about the image for James Watson and Crick?
Some of the X-rays were deflected as they passed through aligned DNA fibers which produced the smudges.
This showed the helical shape of the DNA molecule, also showed that it was made of of two-strands, making a double helix.
This also allowed watson to figure out that the width of the helix was uniform, which also meant the spacing of the nitrogenous bases was uniform, meaning the pairs of bases were uniform.
This allowed Watson to use Chargaffs rules to determine the specificity of the base pairs.
Which bases are purines?
Which are pyridimines?
Purine - Adenine and Guanine
Pyridimine - Thymine and Cytosine
Using her X-ray image of a DNA molecule, what was Franklin able to conclude about the DNA molecule that Watson and Crick used for their work?
She concluded that each strand of DNA werre antiparallel.
and
The backbones of each strands were the sugar-phosphates
and
The bases were paired in the interior of the double-helix
Watson and Crick used this information along with Chargaffs rules to determine the specificity of the base pairs.
IN SHORT, Franklin did what?
X-ray image of DNA
Double helix shape, sugar-phosphate backbone with base paits in interior.
What was the ultimate product of Watson and Cricks work? Did this model limit nucleotide sequences?
The Watson-Crick model of DNA, suggesting a double-helix made of two anti-parallel strands of DNA with specific base pairs using Chargaffs rule.
Though the pairings were limited, the sequences of nucleotides along each separate DNA strand could be configured in countless ways. So no.
Is the Watson-Crick model still used today?
YES, this is a symbol of molecular biology.
They won the nobel prize for their work, Franklin had died before she was eligible for the prize.
What was an important implication that the Watson-Crick model of DNA suggested?
A mechanism for replication.
How did the Watson-Crick model suggestasic a replication mechanism? What would the basic process look like?
It suggested that since the two strandas of DNA are complementary, they can each be used as a template for making a new strand during replication.
Refer to the picture, but the basic mechanism would be that the parent molecule would break into the two strands, and daughter-strands would be build based on the rules of base-pairing.
Describe the watson-crick model of replication, otherwise known as the semiconservative model. Describe the other two competing models of replication known as the conservative model and the dispersive model.
Semi-conservative - when a DNA molecule replicates, each daughter cell will have one old strand of DNA (conserved from parent molecule) and one new strand of DNA.
Conservative - The two parental strands come back together after acting as template for new strands, making the same parental double helix.
Dispersive model - Each daughter strand contains a mixture of parent and newly made DNA
Describe the meselson and stahl experiment that ended up supporting the semi-conservative model of DNA replication. What bacteria was used? What isotopes were used? What was the significance of the isotopes in how they could be differentiated? What did the first replication show? Second replication?
- E. coli were cultured using a medium with N15, a heavier isotope.
- These bacteria were then transferred to a medium contianing only N14, a lighter isotope.
- Bacteria were extracted after their first, and then their second replication, they then were centrifuged
- The purpose of the different isotopes are the label the parent strands (N15) and the new strands formed with the new medium (N14).
- First replication: Both DNA molecules had N15/N14 hybrid, showing some of parent strand in both. This disproves the conservative model that should show separate N15 and N14 DNA strands.
- Second replication: Both N14 only DNA and Hybrid N15/N14 DNA produced. This disproves the dispersive model that should be a mixture of parent and new strand.
Understand this band pattern picture for test that is based on the results of the meselson-stahl experiment.
As the Bacterial DNA replicates more, the amount of N15 labeled decreases in proportion to the N14 labeled DNA strands.
How many enzymes and other proteins participate in DNA replication?
More than a dozen.
Where does DNA replication begin, regardless if they are prokaryotes or eukaryotes?
Origin of replication
Prokaryotes have one origin of replication while eukaryotes have hundreds to thousands.
Describe what an origin of replication is.
This is where the two strands of DNA separate and open up a “replication bubble”.
Look at this picture of DNA replication in a prokaryote and understand it.
DO IT
Understand this basic picture of replication forks of a eukaryote.
DO IT.
Does the antiparallel structure of DNA affect the direction of tis replication? Why?
YES
DNA polymerase only adds nucleotides to the 3’ end of the growing strand, meaning it will only replicate in the 5’ to 3’ direction.
Describe what happens chemically as a nucleotide is added to a grwoing DNA strand.
DNA polymerase catalyzes the addition of the nucleotide to the 3’ end of the growing strand and two phosphate groups are released in the process.
Which strand dictates the overall direction of replication?
The leading strand (it leads the direction)
What do the lagging strand and the leading strand do at each half of the replication bubble?
They shift positions.
Look at this picture and understand what is going on.
DO IT.
What is another repairing mechanism besides mismatch repair that fixes damaged or mismatched nucleotide pairs? Describe it.
Nucleotide Excision Repair - a nuclease (enzyme) is used to excise (cut out) the damaged or mismatched pair, careating a gap. The undamaged strand is used as a template for the correct pairing to occur.
In eukaryotes, because they have linear DNA, what is the limitation of DNA polymerase when it comes to replication of DNA?
What does this limitation result in?
It has to have a 3’ end available in order to attach nucleotides.
At the 5’ end of daughter strand, the RNA primer of the Okazaki fragment on the end cant be replaced with DNA by DNA polymerase. This leaves an overhang of the parent 3’ portion and a shorter daughter strand at its 5’ end. Creating a staggered look.
This means the daughter DNA fragments will get shorter and shorter after each replication.
Describe a nucleosome or a “bead on a string”. What does a nucleosome consist of? What does a protein core consist of? What is linker DNA?
This is completely unfolded chromatin.
Each “bead” is a nucleosome and is 10-nm in diameter
each nucleosome consists of DNA wound around a protein core.
Protein core: 2 molecules of each of the 4 types of histone (H2A, H2B, H3 and H4)
Linker DNA is the DNA wrapped around the middle of the “bead”
Describe a 30-nm fiber.
H1, a 5th histone attaches close to each nucleosome.
Histone tails of nucleosomes interact with other histone tails of other nucleosomes and linker DNA on either side, forming the next level of packing.
Describe looped domains (300-nm fiber).
The 30-nm fiber forms loops as it attaches to a chromosome scaffold made of proteins.
THESE PROTEINS ARE NOT HISTONES
Describe a metaphase chromosome.
The looped domains coil and foil in a manner tht isn’t fully understood yet, but particular genes always end up located in the same places, showing highly precise process.
Width of one chromatid is 700-nm.
