DNA Replication Flashcards
What is cDNA?
DNA synthesised from single stranded RNA (e.g. mRNA or micro RNA) template in a reaction catalysed by the enzyme reverse transcriptase. cDNA is often used to clone eukaryotic genes in prokaryotes
What is centrifugation?
A process that is used for the separation of fluids, gas or liquid based on density
What is conservative replication?
A type of replication that results in one of the daughter molecules containing all the parent DNA and the other daughter molecule containing none of it
What is dispersive replication?
A type of replication that results in all off the daughter molecules having segments of DNA from their parent and the rest is new DNA
What is DNA cloning?
(1) the act of making may identical copies of a DNA molecule - the amplification of a particular DNA sequence. (2) also, the isolation of a particular stretch of DNA from the rest of the cells genome
What is a DNA library?
Collection of cloned DNA molecules representing either an entire genome (genomic library) or complementary DNA copies of the mRNA produced by a cell (cDNA library)
What is DNA polymerase?
An enzyme that synthesises DNA by joining nucleotides together using a DNA template as a guide
What is gel electrophoresis?
A laboratory method used to seperate mixtures of DNA, RNA, or proteins according to molecular size
What is a genomic library?
Collect of cloned DNA molecules representing an entire genome
What is helicase?
A type of enzyme that separates double-stranded DNA into single strands allowing each strand to be copied
What is hybridisation?
The process whereby two complementary nucleic acid strands forms a base paired duplex DNA-DNA, DNA-RNA, or RNA-RNA molecule. Forms the basis of a powerful technique for detecting specific nucleotide sequences
What is ligase?
A specific type of enzyme that facilitates the joining of DNA strands together by catalysing the formation of a phosphodiester bond
What is a plasmid vector?
Small, circular molecules of double stranded DNA derived from plasmids that occur naturally in bacterial cells; widely used for gene cloning
What is a poly a tail?
A stretch of RNA that has only adenine bases
What is polymerase chain reaction?
Technique for amplifying specific regions of DNA by the user sequence specific primers and multiple cycles of DNA synthesis, each cycle being followed by a brief heat treatment to separate complementary strands
What is primase?
An enzyme that synthesises a short strand of RNA on a DNA template, producing a primer for DNA synthesis
What is a primer?
A short strand of RNA that serves as a starting point for DNA synthesis
What is recombinant DNA technology?
Collection of techniques by which DNA segments from different sources are combined to make new DNA, often called recombinant DNA. Recombinant DNAs are widely used in the cloning of genes, in the genetic modification of organisms, and in the production of large amounts of rare proteins
What is replication cell senescence?
Phenomenon observed in primary cell cultures in which cell proliferation slows down and irreversibly halts
What are restriction nucleases?
One of the large number of nucleases that can cleave a DNA molecule at any site where a specific short sequence of nucleotides occurs. Extensively used in recombinant DNA technology
What is semi-conservative replication?
A type of replication that results in both daughter molecules having a parent strand of DNA and a new strand. DNA synthesis is semi-conservative
What is shelterin?
A protein complex known to protect telomeres in many eukaryotes from DNA repair mechanisms, as well as regulate telomerase activity
What are single-stand binding proteins?
A protein that binds to the single strands of the opened up DNA helix, preventing helical structures from reforming while the DNA is being replicated
What are somatic cells?
Any cell of a plant or animal other than the cells of the germ line
What is strand-directly mismatch repair?
A proofreading system that removes DNA replication errors missed by the DNA polymerase proofreading exonuclease. It detects the potential for DNA helix distortion from noncomplememtary base pairs the recognises the excises the mismatch in the newly synthesised strand and resynthesises the excised segment using the old strand as a template
What is taq polymerase?
An enzyme found in the bacillus Thermus aquaticus, which lives in hot springs; it is heat resistant and thus can endure the high temperatures of the polymerase chain reaction
What are tautomeric forms?
Constitutional isomers of the main forms of nucleotides that may cause mistakes in base pairing
What is telomerase?
Enzyme that elongates telomere sequences in DNA, which occurs at the ends of eukaryotic chromosomes
What is a telomere?
End of a chromosome, associated with a characteristic DNA sequence that is replicated in a special way. Counteracts the tendency of the chromosome to otherwise shorter. With each round of replication
How many origins of replications occur in prokaryotes?
One
How many origins of replication occur in eukaryotes?
Multiple
What enzyme opens up the DNA?
Helicase
What is responsible for stabilising unwound DNA?
Single-strand binding protein
Which enzyme is responsible for adding nucleotides to the growing polynucleotide chain? What direction does it synthesise in?
DNA polymerase
5’ to 3’
How do enzymes know where to start replicating the DNA?
The replication sites have short sequences to attract the initiator proteins and stretches of DNA that are easy to open
Is eukaryotic or prokaryotic replication quicker?
Prokaryotic
Replication requires RNA primers, what enzyme(s) add these primers to the DNA?
DNA primase
How are old primers removed from the DNA?
They are erased by a combined action of DNA polymerase and RNase H
What does the clamp loader do?
Holds the DNA polymerase onto the DNA as it moves
Which enzyme is responsible for relieving helical winding?
DNA topoisomerases
How often does a mistake occur in DNA replication?
1 in a billion
In the rare tautomeric form, what base can C pair with?
A instead of G
After covalent binding, what is the next error correcting reaction?
Exonucleolytic proofreading by DNA polymerase in the 3’ to 5’ direction
How does exonucleolytic prrofreading work?
Correct nucleotide pairing has a higher affinity for polymerase. If an incorrect pairing occurs, the polymerase chews back to create a base-paired 3’ -OH end on the primer strand. It then continues the process of adding nucleotides correctly
What would happen if the strand-directed mismatch repair system was not able to differentiate between the new strand and old strand?
It would correct the wrong nucleotide 50% of the time
What allows the strand-directed mismatch repair system to differentiate between the new strand an the old strand?
Newly synthesised lagging-strand DNA transiently contains nicks (that haven’t yet been sealed by DNA ligase) and such nicks provide the signal that directs the mismatchprrofreading to the appropriate strand
What enzyme adds telomere sequences to the end of chromosomes?
Telomerase
What is the proteins that form a protective chromosome cap called?
Shelterin
What does shelterin do?
Shelterin hides telomeres from the cell’s damage detectors that continually monitor DNA
What happens when there is no telomere sequence left?
The cell enters replicative cell senescence, withdrawing them permanently from the cell cycle and preventing them from dividing
