Lecture 2 Flashcards
What were the key experiments that led to the conclusion that DNA is the carrier of genetic information?
Griffiths
Avery, MacLeod, & McCarty
Hershey & Chase
Watson & Crick
When was the griffiths experiment
1928
What did the griffiths experiment do?
Discover the Transforming Principle
Describe the bacteria Streptococcus pneumonia used in the Griffiths experiment
Has 2 forms: rough and smooth
Smooth has a protective polysaccharide layer called glycocalyx, is virulent
Rough doesn’t have glycocalyx, isn’t virulent
Why is the smooth form of Streptococcus virulent?
The glycocalyx protects the bacteria from the mouse’s immune system
What happened when the smooth form of Streptococcus was heat treated?
The heat killed the s strain, and was no longer virulent
No s strain could be obtained from mice after injection
What happened when heat treated s-strain of Streptococcus was injected along with the r strain?
The mouse died
Live R and S-strains could be obtained from the mice after injection
What was the conclusion of the griffiths experiment
living cells (“R”) could be “transformed” by dead (“S”) cells Something in the S-cell debris could convert (transform) R cells into S cells
T/F certain bacteria can take up fragments of DNA
True
What do bacteria need to be in order to take up dna?
Need to be competent
What is bacteria competency caused by?
Competency factors
What are competency factors?
Single stranded binding proteins, which help incorporating the ingested DNA into the bacterial genome
When was the Avery, MacLeod & McCarty experiment
1944
What did Avery, MacLeod & McCarty do?
Treated S-strain with compounds that would destroy different elements of the bacteria
What were the elements that were destroyed in the Avery, MacLeod & McCarty experiment?
Glycocalyx Lipids RNA Protein DNA
What happened after each element was destroyed?
When glycocalyx, lipids, RNA, and protein were destroyed, the activity stayed
But when DNA was destroyed, activity was lost
What was the conclusion of the Avery, MacLeod & McCarty experiment
DNA is the transforming substance
T/F Avery, MacLeod & McCarty’s results were widely accepted
False
People were reluctant to accept DNA was the genetic material
why were people reluctant to accept dna as the genetic material?
DNA seemed not complex enough (compared to proteins) to be the base for genetic material (4 different nucleotides vs. 20 amino acids).
When was the Hershey & Chase experiment?
1952
Describe the bacteriophage T2 used in the Hershey & Chase experiment
It is composed of only DNA and protein, therefore the genetic material had to be either protein or DNA
What did Hershey & Chase do with the bacteriophages?
Labelled some with 32P to follow DNA
and some with 35S to follow Protein
What were the results of Hershey & Chase experiment when they labelled bacteriophage with 35S?
35S labels proteins
Most of the radioactivity was recovered in phage ghosts
What were the results of Hershey & Chase experiment when they labelled bacteriophage with 32P?
32P labels DNA
Most of radioactivity was recovered inside the bacteria
Explain the results of the Hershey & Chase experiment, in terms of what was going on with the bacteriophage
Upon injection, only the dna enters the bacterium, the empty phage “ghosts” remain outside the bacterium
What was the conclusion of the Hershey & Chase experiment
DNA is the hereditary material (NOT protein). At least for bacteriophage T2…
What realizations did we have after the Hershey & Chase experiment
Cells must be able to replicate DNA. Carries hereditary information Transfer information to control a cell's activity Must be able to change (mutate). How does this all happen?
Describe nucleotide structure
Phosphate group
Pentose sugar
Nitrogenous base
How do we name nucleotides?
Base + sugar + phosphate
How do we name nucleosides
Base + sugar
How are nucleotides linked?
Phosphodiester linkage
Phosphate group at 5’ of the sugar of one nucleotide covalently bonds to the 3’ of the sugar on the next nucleotide
T/F the phosphate backbone of DNA is positively charged
False
Negative charged
What did Chargaff do in the 1940’s
He looked at the nucleotide distribution across species
What did chargaff find?
The chemical composition of dna across species is the same
That each species has a specific quantity of dna
Dna found in different species have the same ratio of bases
What are chargaff’s rules?
A = T
C = G
In rna, A = U
What did rosalind franklin do
Produce an X-ray diffraction image of DNA
How are nitrogenous bases connected to each other?
Hydrogen bonds
Minor groove
The backbones are close together
Major groove
The backbones are further apart
Describe dna structure
2nm wide Helical 2 strands run antiparallel Sugar and phosphate face out, bases face in 10.5 bases per helical turn
Purine
Double ring
A and G
Pyrimidine
Single ring
T and C and U
How did we conclude that pyrimidine pairs with purine?
Spacing, pyra + pyra = too narrow, pur + pur = too wide
Pyra + pur = consistent with xray data
Complementary base pairing
A always pairs with T (or U)
C always pairs with G
What are the three models of DNA replication models?
Conservative
Semiconservative
Dispersive
Describe the conservative model of dna replication
Parent strands separate
Each parental strand is replicated, creating a two DNA duplexes, each with one parental and one daughter strand
These new strands separate
The parental strands reanneal and the daughter strands anneal
If the conservative model of dna replication is true, how much new vs. old DNA is present in first round of replication and in second round of replication?
First: one duplex is 100% old, one duplex is 100% new
Second: one duplex is 100% old, three duplexes are 100% new
If the conservative model of dna replication is true, what is the ratio of old vs. new DNA in first round of replication and in second round of replication?
First: 1:1
Second: 1:3
Describe the semiconservative model of dna replication
Parent strands separate
Each parental strand is replicated, creating a two DNA duplexes, each with one parental and one daughter strand
If the semiconservative model of dna replication is true, how much new vs. old DNA is present in first round of replication and in second round of replication?
First: both duplexes are 50% old and 50% new
Second: two duplexes are 50% old and 50% new, two duplexes are 100% new
If the semiconservative model of dna replication is true, what is the ratio of old vs. new DNA in first round of replication and in second round of replication?
First: 1:1
Second: 1:3
Describe the dispersive model of dna replication
Segments of parental DNA and newly synthesized daughter DNA are interspersed amongst both strands of the DNA molecules
If the dispersive model of dna replication is true, how much new vs. old DNA is present in first round of replication and in second round of replication?
It would vary, but there would be around 50% new dna after each round of replication
Which replication model is correct
Semiconservative
How did we figure out which model of dna replication was correct?
Through the Meselson-stahl pulse-chase experiment
What happened in the first part of the Meselson-stahl pulse-chase experiment
Grew bacteria in a medium with heavy 14N, which would be incorporated in the dna
After enough time that the isotope has been incorporated, but before dna is replicated, the bacteria is transferred to fresh medium without the isotope, so new dna would not have the isotope
What happened in the second part of the Meselson-stahl pulse-chase experiment
The dna was put in CsCl solution and spun really fast to make a gradient where heavier dna was at the end
T/F for the Meselson-stahl pulse-chase experiment, they ran it for 3 rep cycles
False
Two rep cycles
In the Meselson-stahl pulse-chase experiment, why did they run it for 2 cycles?
Because after one rep cycle, semiconservative and dispersive both have 50% of parent dna, they’re both in the middle of the solution
