Complete, clear and concise notes for Topic 7-11, the HL portion of the IB Biology course, by a 44 student (7 for biology) student
Notes are structured according to the syllabus requirements to ensure all necessary information is covered, and includes all needed diagrams and possible long answers
Requirements from the official IB Biology guide; important points; definitions/equations; not needed
Topic 7: Nucleic Acids
7.1 DNA structure and replication
Understandings:
● Nucleosomes help to supercoil the DNA.
● DNA structure suggested a mechanism for DNA replication.
● DNA polymerases can only add nucleotides to the 3’ end of a primer.
● DNA replication is continuous on the leading strand and discontinuous on the lagging strand.
● DNA replication is carried out by a complex system of enzymes.
● Some regions of DNA do not code for proteins but have other important functions.
Applications:
● Rosalind Franklin and Maurice Wilkins’ investigation of DNA structure by X-ray diffraction.
● Use of nucleotides containing dideoxyribonucleic acid to stop DNA replication in preparation of samples for base
sequencing.
● Tandem repeats are used in DNA profiling.
Skills:
● Analysis of results of the Hershey and Chase experiment providing evidence that DNA is the genetic material.
● Utilisation of molecular visualisation software to analyse the association between protein and DNA within a nucleosome.
Hershey-chase Experiment
Analysis of results of the Hershey and Chase experiment providing evidence that DNA is the genetic material.
Hersey-chase: Determine whether proteins or DNA is passed down from parents becomes genetic material of the cell
- While prokaryotic DNA is naked DNA, Eukaryotic chromosomes are associated with proteins
- Sperm and egg cells contain both DNA and proteins within the nucleus (as does the zygote)
- Confusion over whether it is the proteins or DNA that allow for inheritance of hereditary characteristics
Investigation using a virus (T2) that infects E. coli (gut bacteria)
(Alfred Hershey & Martha Chase)
- Known that when viruses infect, they pass on genetic material - viral proteins made in bacteria
- DNA contains phosphorus (phosphate group), Protein coat contains sulphur (in amino acids methionine and
cysteine)
Autoradiography - radioactively labelling allowing for future detection
- Viruses (T2 bacteriophage) grown in isotopic mediums to radioactively label a specific viral component
- Virus grown in radioactive sulphur to label DNA with radioactive 32P
- Another virus grown in radioactive phosphorus to label protein with radioactive 35S
- The viruses are then mixed with the E.coli, infecting them
Centrifugation - separate contents according to density using centripetal force
- Using centrifugation, mixture is separated into a pellet (bottom) containing the heavier bacteria and liquid
supernatant (top) containing the lighter viruses
- The radioactive 32P (DNA) was mostly found in the pellet (containing bacteria) while the radioactive 35S (protein)
was only found in the supernatant (containing viruses)
Results - Thus, it can be determined that the DNA is what is injected into the bacteria as only it is found in the pellet
- Strong evidence that genes are composed of DNA rather than proteins - DNA is genetic
Franklin and Wilkins - DNA & X-ray diffraction
Rosalind Franklin and Maurice Wilkins’ investigation of DNA structure by X-ray diffraction.
Used to prove the double helix structure of DNA - known that DNA is double stranded, but actual structure unknown
Experiment by Rosalind Franklin and Maurice Wilkins
Three aspects to her investigation
, 1. Use of X-ray diffraction to identify structure
Visible light - 400-700 nm, cannot be used to study DNA structure as DNA width is smaller than the vl wavelength
X-ray - 0.1 - 10 nm, give high resolution image of the DNA molecule (lower wavelength → higher resolution)
2. Use crystalline sample of DNA
Only solid objects can diffract x-rays - as beam of x-ray passes through sample, it is diffracted by the solids
3. Photographed DNA in 3 cartesian coordinates (x, y, z)
Rotating the sample in three different dimensions - verify helical orientation in 3D
The diffraction patterns were recorded using x-ray films
Results
- Confirmed that Dna has a helical structure
- Phosphate groups were on the outside making use of the X-ray diffraction pattern (phosphate-sugar backbone)
- The distance between each bend in the helix is 3.4 nm (34 Angstroms)
- Each bend contains 10 base pairs, so each base pair has width of 0.34 nm (3.4 A)
DNA structure suggested a mechanism for DNA replication.
- Replication occurs by complementary base pairing (adenine with thymine, guanine with cytosine)
- Replication is bi-directional (proceeds in opposite directions on the two strands) due to the antiparallel nature
Nucleosome & Supercoiling
Nucleosomes help to supercoil the DNA.
DNA molecules stretch out to ~2m, but is able to be stored in a few nm within the nucleus due to supercoiling of DNA
- DNA in eukaryotes are associated with histone proteins - which play a vital role in condensing the DNA
Nucleosome - a DNA molecule wrapped around 8 histone proteins and a H1 histone protein
Condensed DNA
- Negatively charged DNA (from phosphate groups) attracted to positively charged histone proteins
- This allows DNA to wrap twice around the 8 histone proteins (protein core)
- And around linker protein (histone protein H1) which binds to outside of DNA strand
Supercoiling
Analyse the association between protein and DNA within a nucleosome.
- H1 protein have N-terminal tails sticking outwards from the nucleosomes
- N-terminal ends of histone complexes can undergo further condensation reaction - causing DNA to coil further
- During uncoiling, it gives the entire DNA the appearance of string of beads - beads are nucleosomes, chromatin
is the long string of beads
Structure of nucleus complexes regulates
1. Mitosis
- Conversion of chromatin to X-shaped chromosomes (nucleosome arrangement)
2. Gene Expression
- Regulates transcription
- Different parameters allows access of certain areas of DNA (switch on gene) for transcription (for
production of mRNA)
- If nucleosomes do not open up, there is no gene expression
Organisation of Eukaryotic DNA
- DNA is complexed with eight histone proteins (an octamer) to form a complex called a nucleosome
- Nucleosomes are linked by an additional histone protein (H1 histone) to form a string of chromatosomes
- These then coil to form a solenoid structure (~6 chromatosomes per turn), which condense to form a 30 nm
fibre
- These fibres then form loops, which are compressed and folded around a protein scaffold to form chromatin
- Chromatin will then supercoil during cell division to form chromosomes that are visible under microscope
DNA replication
Replication - formation of a new DNA molecule: occurs before a cell divides during interphase (S phase of cell cycle)
DNA replication is semi conservative (Meselson and Stahl)
DNA replication (semi conservative) (7-9 Long mark answer)
, - During the S phase of the cell cycle, DNA is semi-conservatively replicated into two identical copies
- In eukaryotic cells, the DNA is very long, therefore replication occurs at many different regions on the DNA
strand called the Origins of replication
- Replication takes place in 5’ to 3’ direction but oppositely for the two strands as they are anti-parallel
- First, DNA Helicase breaks H bonds between nitrogenous bases to unwind DNA into two strands at the Origin
- There are multiple Origin of replications, with the replication fork at each end of where the DNA unwinds
- The strain from unwinding the two strands are released and stabilised by DNA Gyrase and stabilising proteins
- Single stranded binding (SSB) proteins attach to the strands to keep DNA from winding back
- DNA primase inserts RNA primers to the two strands of DNA to initiate and indicate place for start of replication
(one primer to the strand 5’ to 3’ and multiple primers to the strand 3’ to 5’)
- DNA polymerase III adds free DNA nucleotides, which exists as nucleoside triphosphates in the 5’ to 3’ direction
following complementary base pairing (A-T 2 H bonds, C-G 3 H bonds)
- Nucleoside triphosphate - nucleotides with three phosphate groups instead of one, during binding DNA
polymerase cleaves off the two phosphate groups, releasing energy to form the phosphodiester bond for
sugar-phosphate backbone
- DNA polymerase I removes the RNA primers and replaces it with DNA nucleotides (triphosphates)
- DNA replication is continuous on the leading strand and discontinuous on the lagging strand.
- The strand with one primer is called the leading strand and the one with multiple is called the lagging strand
- Replication on the leading strand (5’ to 3’) is continuous as it is moving towards the replication fork
- Replication on the lagging strand (3’ to 5’) is discontinuous as it is moving away from the replication fork
- DNA polymerases can only add nucleotides to the 3’ end of a primer.
- Polymerase can only add nucleotides in 5’ to 3’ direction, so replication of lagging strand results in series
of fragments known as Okazaki fragments
- DNA Ligase sticks the Okazaki fragments together in the lagging strand to make it continuous
- Thus two identical strands of DNA molecule is formed - each molecule containing half the original DNA material
Enzymes in DNA replication (6 Enzymes)
DNA replication is carried out by a complex system of enzymes.
Enzyme Role in DNA replication
DNA Helicase unwinds the DNA by breaking H bonds breaking nitrogenous bases
DNA Gyrase (and single stabilises DNA strand by releasing strain developed within the strand
strand binding proteins)
DNA Primase synthesises RNA primers and attaches them to the two strands (primers attach to one
strand only)
DNA Polymerase III adds nucleoside triphosphates to the DNA strand
DNA Polymerase I removes the primers on the strand and replaces the gaps with nucleotides
DNA Ligase Joins the okazaki fragments together by forming phosphodiester bonds
DNA base sequences
Genes within the DNA are called coding sequences - the order of bases in DNA code for polypeptides (proteins) created
during transcription and translation
However, only less than 2% of DNA are for coding sequences
These are known as Exons - useful DNA in production of proteins and hereditary characteristics (Introns are non-coding
sequences)
The remaining 98% of DNA is used for other functions (non-coding sequences) or have no function
Some regions of DNA do not code for proteins but have other important functions.
, The regions of DNA that do not code for proteins should be limited to regulators of gene expression, introns, telomeres
and genes for tRNAs.
Functions of Non-coding sequences/Introns (STINGS)
1. Satellite DNA
- Long stretches of DNA made up of highly repetitive sequences - short tandem repeats (STRs)
- Structural component of heterochromatin and centromeres
- Used for DNA profiling/fingerprinting - as they are unique to each person (like fingerprints)
- Identification of individuals (forensic studies) and paternity testing
2. Telomeres
- Regions of repetitive DNA at the end of a chromosome
- Protects against chromosomal deterioration during replication, acts as buffer to strain DNA is subjected
to - ageing happens when telomeres run out as DNA starts to be lost/deteriorate
- Hydras - biologically immortal (does not age) as their telomeres does not shorten
- Lobsters - have telomerase, can continue repairing telomeres
- Age can be identified by length of telomere
3. Introns
- Non-coding sequences within genes
- Removed by RNA splicing prior to formation of mRNA (before leaving the nucleus)
- Includes Satellite DNA and all other non-coding sequences
4. Non-coding RNA genes
- Codes for RNA molecules that are not translated into protein (only transcripted)
- Production of tRNA (transfer) and rRNA (ribosomal)
5. Gene regulatory sequences
- Sequences involved in the process of transcription - includes promoters, enhancers and silencers
- The promoter as an example of non-coding DNA with a function.
- Promoters - sequence indicating the start site for transcription, which RNA polymerase binds to
during transcription initiation
- Enhancers - sequences that regulatory proteins bind to to enhance gene expression
- Silencers - sequences that regulatory proteins bind to to inhibit gene expression (by preventing
RNA polymerase from binding to promoter site)
- Regulatory sequences are transcribed
6. Structural DNA
- Highly coiled DNA with no coding function
- Helps keep structure of DNA
- Near centromere and telomere
DNA Sequencing and Profiling
Use of nucleotides containing dideoxyribonucleic acid to stop DNA replication in preparation of samples for base
sequencing.
Process that explains the base order of a nucleotide sequence - sequencing of the genome
Sanger based sequencing
- Uses dideoxynucleotides (ddNTPs) - instead of OH on second and third C, it has only H (C5H10O3)
- DNA polymerase can only add nucleotides to the 3’ ends of a primer
- The OH on the third C is vital for formation of sugar-phosphate backbone - thus ddNTPs prevents further
elongation of the DNA strand and terminates replication when it is added to a new DNA strand
- Fluorescent dye markers are attached to dDNA so that the base present when replication stops can be identified
- This creates different sized fragments with fluorescent markers that can be separated by gel electrophoresis by
length of strand
- Sequences of bases analysed by comparing the colour of the fluorescent fragment (the terminal base)
Tandem repeats are used in DNA profiling.
Profiling - use of Short Tandem Repeats (STRs) in DNA to identify people
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