Topic 7: Nucleic Acids Flashcards
(35 cards)
Outline how Franklin and Wilkins used X-ray crystallography to elucidate the structure of DNA
DNA was crystallised and then targeted with an X-ray (whose beam became diffracted by DNA crystals). The scattering pattern created by the diffracted X-ray was recorded on film. This pattern was then analysed to elucidate the structure of DNA.
Describe the organisation of DNA into chromatin within eukaryotic cells
DNA is wrapped around histone proteins to form nucleosomes. Nucleosomes are grouped together (chromatosomes) and then arranged into fibres (chromatin). DNA is usually organised as chromatin within the nucleus, except during cell division (when the chromatin condenses to form chromosomes).
Differentiate between euchromatin and heterochromatin
Euchromatin is more loosely packed and corresponds to active segments of DNA (i.e. active genes). Heterochromatin is more densely packaged and corresponds to inactive segments of DNA. Different cells have different segments of DNA packaged as euchromatin and heterochromatin.
Outline the structure of the nucleosome (and identify its functions)
A nucleosome consists of DNA and histone proteins. DNA is wrapped around an octamer of histone proteins. Nucleosomes are linked by an interconnecting H1 histone.
Nucleosomes serve two key functions: they help to supercoil DNA, they help to regulate transcription.
List five examples of non-coding DNA
Satellite DNA (e.g. short tandem repeats), telomeres, introns, non-coding genes, gene regulatory sequences.
Explain the role of tandem repeats in DNA profiling
Short tandem repeats are short repeating segments within satellite DNA. The number of repeats for a particular loci will differ between individuals. The STRs can be excised and separated on a gel to create a distinct DNA profile of a given individual.
Outline the methodology and conclusions of the Hershey-Chase experiment
Hershey and Chase demonstrated that DNA was the genetic material by using radioactively labelled viruses. Viruses were prepared with radioactive phosphorus (labels DNA) or radioactive sulphur (labels protein). Viruses then infected bacteria, before the bacteria and virus were separated via centrifugation (bacteria is heavier and forms a pellet, while the smaller virus remains in the supernatant). When radioactive sulphur was used, radioactivity was detected in supernatant (not transferred to bacteria). When radioactive phosphorus was used, radioactivity was detected in pellet (WAS transferred to bacteria)
State the direction of DNA replication
Replication occurs in a 5’ to 3’ direction (on the newly synthesised strand)
State the role of the following enzymes of the DNA replication process
Helicase: unwinds and separated double stranded DNA
DNA gyrase: relieves torsional strain created by helicase action to prevent supercoiling
SSB proteins: prevents DNA strands from re-annealing
DNA primase: lays down a short RNA primer to provide an initiation point for polymerisation (DNA pol III can only add nucelotides to the 3’-end of an existing nucleotide chain)
DNA polymerase III: extends the nucleotide chain from the primer
DNA polymerase I: removes and replaced RNA primers with DNA nucleotides
DNA ligase: covalently joins Okazaki fragments together (on lagging strand)
Outline the difference between leading and lagging strands as the relate to Okazaki fragments
DNA strands are antiparallel, so DNA polymerase III moves in opposite directions on the two strands. On the leading strand, DNA polymerase III moves in the same direction as helicase so synthesis continuous. On the lagging strand, DNA polymerase III moves in the opposite direction to helicase - synthesis is discontinuous. The fragments generated on the lagging strand are called Okazaki fragments.
Describe the role of deoxynucleoside triphosphates (dNTPs) in the replication process
Deoxynucleoside triphosphates (dNTPs) align opposite their complementary base partner. DNA pol III cleaves two of the phosphates and uses the energy to form a covalent phosphodiester bond. In this way, DNA pol III synthesises a new DNA strand
Explain how dideoxynucleotides are used in DNA sequencing
Dideoxynucleotides (ddNTPs) lack the 3’-hydroxyl group needed to form a phosphodiester bond. This means the inclusion of a ddNTP will terminate the extension of a DNA sequence at that point. Four PCR cycles are set up, each with a different ddNTP (ddA, ddT, ddG or ddC) and a stock of normal bases. Each time the ddNTP is incorporated the sequence stops, generating fragments. When these fragments are separated and then ordered according to length, the DNA sequence is discerned. This process can be automated by using fluorescently labelled ddNTPs that can be detected by machine.
Distinguish between the sense and antisense strands
The antisense strand is transcribed into mRNA (complementary to the DNA template). The sense strand is not transcribed into mRNA (it is identical to the DNA template.
Identify the three sections of a gene
Promoter - initiation point (where RNA polymerase binds)
Coding Region - sequence that is transcribed
Terminator - termination point
Explain the process of transcription including the role of RNA polymerase
RNA polymerase attaches to the promoter (requires the presence of transcription factors)
RNA polymerase unwinds the DNA and separates the strands
Nucleoside triphosphates (NTPs) align opposite complementary bases and are joined by RNA polymerase
This continues until the terminator, at which point the enzyme and transcript dissociate from the DNA
List three examples of post-transcriptional modifications
Methyl cap added to the 5’-end (protects against degradation and allows recognition by ribosome)
Polyadenylation occurs at the 3’-end (improves stability and facilitates nuclear export)
Splicing occurs (introns are removed)
Distinguish between introns and exons
Introns are intruding sequences within a gene that are removed prior to translation (not expressed). Exons are the expressing sequences translated into protein.
Outline how alternative splicing increases the number of different proteins an organism can produce
Exons can be selectively removed via alternative splicing to produce different protein variants from the same gene - this increases the number of proteins an organism can produce relative to the number of genes
Outline the role of the promoter and transcription factors in the regulation of transcription
Transcriptional activity is regulated by transcription factors which mediate binding of RNA polymerase to the promoter (initiation point for transcription of a gene sequence). The presence of certain transcription factors may be tissue-specific (explaining why different genes are activated in different tissues) and environmentally sensitive (explaining why expression patterns can change).
Discuss the role the environment can play in regulating transcription
Environmental factors can trigger the production of transcription factors. Example, hydrangea flowers change colour according to the pH of the soil.
Describe the role regulatory proteins play in moderating gene expression levels
The binding of RNA polymerase to the promoter can be controlled by regulatory proteins. Activators bind to enhancer sequences and make it easier for the enzyme to bind (increases expression). Repressors bind to silencer sequences and make it harder for the enzyme to bind (decreases expression).
Outline the role of the nucleosome in the regulation of gene expression
Nucleosomes are involved in the supercoiling of DNA (supercoiled DNA is accessible for transcription). Acetylation of the histones makes the DNA less tightly coiled (increased gene expression). Methylation of the histones makes the DNA more tightly coiled (decreases gene expression).
Describe how DNA methylation patterns may differ between identical twins and the significance of this
Direct methylation of DNA also decreases gene expression (prevents binding of transcription factors). Hence genes that are methylated are not expressed, while genes that are not methylated are expressed. Because DNA methylation is influenced by environmental factors, identical twins will develop different methylation patterns over time - this explains why twins may look different despite have identical genomes.
Identify the following sites on a tRNA molecule and describe their function
Acceptor stem - carries the amino acid
T arm - associates with the ribosome (via E, P, A sites)
Anticodon - associates with mRNA (via complementary codon)
D arm - associates with tRNA-activating enzyme
