A1: Neurobiology HL Extension Flashcards
What did Hershey and Chase figure out?
In 1952, Alfred Hershey and Martha Chase conducted a series of experiments to prove that DNA was the genetic material.
Viruses (T2 bacteriophage) were grown in one of two isotopic mediums in order to radioactively label a specific viral component.
Viruses grown in radioactive sulfur (35S) had radiolabelled proteins (sulfur is present in proteins but not DNA)
Viruses grown in radioactive phosphorus (32P) had radiolabeled DNA (phosphorus is present in DNA but not proteins)
The viruses were then allowed to infect a bacterium (E. coli) and then the virus and bacteria were separated via centrifugation.
The larger bacteria formed a solid pellet while the smaller viruses remained in the supernatant.
The bacterial pellet was found to be radioactive when infected by the 32P–viruses (DNA) but not the 35S–viruses (protein).
This demonstrated that DNA, not protein, was the genetic material because DNA was transferred to the bacteria.
What did Rosalind and Franklin figure out?
From the scattering pattern produced by a DNA molecule, certain inferences could be made about its structure
Composition: DNA is a double stranded molecule
Orientation: Nitrogenous bases are closely packed together on the inside and phosphates form an outer backbone
Shape: The DNA molecule twists at regular intervals (every 34 Angstrom) to form a helix (two strands = double helix)
What did the Watson and Crick Model suggest about semi-conservative replication?
● DNA structure suggested a mechanism for DNA replication.
● Pyrimidine paired with purine.
● Bases are upside down in relation to one another as strands are antiparallel.
● Adenine has a surplus positive charge and Thymine has a surplus negative charge.
o They are therefore electrically compatible.
● Cytosine paired with Guanine forms three hydrogen bonds, enhancing
stability.
Role of nucleosomes in DNA packing
● Nucleosomes help to supercoil DNA.
● 2 copies of 4 types of histones linked together by linker DNA.
o Connects one nucleosome to the next.
o H1 (histone number 9) serves to bind DNA to the core particle of 8 histones.
● Process is supercoiling.
o Tails of histone proteins interact with tails from histone proteins in other nucleosomes to tighten and supercoil the chromosome.
● H1 histone bins to form structure called the 30nm fibre that facilitates
further packing.
● Packed together in the nucleosome.
The leading strand and the lagging strand:
● Replication is continuous on the leading strand along the replication fork
as it opens; and discontinuous on the lagging strand moving away from
the replication fork.
● Replication occurs anti-parallel.
● Fragments on the lagging strand are called Okazaki fragments.
What are the proteins that are involved in replication:
● Proteins are involved as enzymes at each stage but also serve several
other functions.
● DNA is unwound by DNA Helicase at the replication fork.
● The strain developing ahead of helicase is released for unwinding by DNA Gyrase (this is a specific class of DNA Topoisomerases).
● The two strands are kept apart by single stranded binding proteins to
allow the template strand to be copied.
● Multiple RNA primers on lagging strand but only one on leading strand.
o Necessary to start replication.
o These are created by DNA primase.
o Initiates activity of DNA polymerase III.
o DNA polymerase I replaces the primer with complementary nucleotides.
● DNA Polymerase III covalently links deoxyribonucleotide-triphosphates
to the 3’ end of the growing strand.
● DNA ligase links Okazaki fragments - specifically, creates sugar phosphate
bonds between the fragments.
What is the direction of replication?
● Replication begins at the origin of replication, where the RNA primer is
placed by DNA primase.
● Replication occurs in the 5’ to 3’ direction (nucleotides are added to the 3’ end of the primer)
Function of non coding DNA?
● Non-coding DNA has some function.
o Regulate gene expression.
o Produce tRNA and rRNA.
o Called introns.
● Repetitive sequences:
o 60% of human genome.
● Telomeres:
o Occur at the ends of eukaryotic chromosomes.
o Serve to protect DNA.
o Genes at the end of chromosomes would be lost if replication continued to the end of a chromosome without a telomere.
o Telomere’s highly repetitive sequences are sacrificed.
Process of DNA profiling
● Variable number tandem repeats (VNTR) distinguishes individuals based
on the number of times this short sequence of DNA repeats.
● Inherited as an allele and is analysed in DNA profiling.
● Sections of VNTR allele combinations are cut from DNA using restriction
enzymes and analysed using gel electrophoresis.
● Two individuals can be compared in this way.
● Paternal lineage can be deduced by analysing alleles of VNTR from the Y
chromosome.
● Maternal lineage can be deduced by analysing variations in mitochondrial
DNA at specific hyper-variable region.
Explain DNA sequencing
● Copies of DNA are placed into test tubes with deoxyribonucleotides and
enzymes for replication.
● Small quantities of fluorescent dideoxyribonucleotides are added which will stop replication upon being added.
o Dideoxyribonucleotides have H instead of OH on the 3rd carbon,
which prevents the binding of a phosphate group to it, hence
stopping the DNA replication process.
● Fragments created will be analysed through gel electrophoresis and
sequences will be determined through colour pattern of fluorescent
markers.
o New technology allows for the fluorescent dideoxyribonucleotides
to be detected by computers and quick interpretation of results.
What is the promotor?
non-coding DNA to which RNA polymerase binds. Not
transcribed.
What is the coding sequence?
After RNA polymerase has bound to the promoter, it causes the DNA strands to unwind and separate
The region of DNA that is transcribed by RNA polymerase is called the coding sequence
What is the terminator?
RNA polymerase will continue to transcribe the DNA until it reaches a terminator sequence
The mechanism for transcriptional termination differs between prokaryotes and eukaryotes
Sense vs antisense strands?
The antisense strand is the strand that is transcribed into RNA
Its sequence is complementary to the RNA sequence and will be the “DNA version” of the tRNA anticodon sequence
The antisense strand is also referred to as the template strand
The sense strand is the strand that is not transcribed into RNA
Its sequence will be the “DNA version” of the RNA sequence (i.e. identical except for T instead of U)
The sense strand is also referred to as the coding strand (because it is a DNA copy of the RNA sequence)
How does transcription occur?
The process of transcription can be divided into three main steps: initiation, elongation and termination
In initiation, RNA polymerase binds to the promoter and causes the unwinding and separating of the DNA strands
Elongation occurs as the RNA polymerase moves along the coding sequence, synthesising RNA in a 5’ → 3’ direction
When RNA polymerase reaches the terminator, both the enzyme and nascent RNA strand detach and the DNA rewinds
Many RNA polymerase enzymes can transcribe a DNA sequence sequentially, producing a large number of transcripts
In eukaryotes, post-transcriptional modification of the RNA sequence is necessary to form mature mRNA
What are the 3 post transcriptional events that must occur in order to form mature messenger RNA?
Capping
Capping involves the addition of a methyl group to the 5’-end of the transcribed RNA
The methylated cap provides protection against degradation by exonucleases
It also allows the transcript to be recognised by the cell’s translational machinery (e.g. nuclear export proteins and ribosome)
Polyadenylation
Polyadenylation describes the addition of a long chain of adenine nucleotides (a poly-A tail) to the 3’-end of the transcript
The poly-A tail improves the stability of the RNA transcript and facilitates its export from the nucleus
Splicing
Within eukaryotic genes are non-coding sequences called introns, which must be removed prior to forming mature mRNA
The coding regions are called exons and these are fused together when introns are removed to form a continuous sequence
Introns are intruding sequences whereas exons are expressing sequences
The process by which introns are removed is called splicing
