2.7 DNA -> Protein Flashcards
What type of process id DNA replication?
DNA replication is a semi-conservative process whereby pre-existing strands act as templates for newly synthesised strands
What two enzymes control DNA replication?
The process of DNA replication is coordinated by two key enzymes – helicase and DNA polymerase
What is the role of helicase?
Helicase unwinds the double helix and separates the two polynucleotide strands
How does helicase unwind the double helix?
It does this by breaking the hydrogen bonds that exist between complementary base pairs
Why is DNA helicase needed?
The two separated polynucleotide strands will act as templates for the synthesis of new complementary strands
What is the role of DNA Polymerase?
DNA polymerase synthesises new strands from the two parental template strands
How do bases align during DNA replication?
Free deoxynucleoside triphosphates (nucleotides with 3 phosphate groups) align opposite their complementary base partner
How does DNA polymerase cause the complementary bases to match with the DNA strand?
DNA polymerase cleaves the two excess phosphates and uses the energy released to link the nucleotide to the new strand
What is PCR?
The polymerase chain reaction (PCR) is an artificial method of replicating DNA under laboratory conditions
Why and when is PCR used?
The PCR technique is used to amplify large quantities of a specific sequence of DNA from an initial minute sample
What 3 conditions does a thermal cycler need for PCR to occur?
Denaturation – DNA sample is heated (~90ºC) to separate the two strands
Annealing – Sample is cooled (~55ºC) to allow primers to anneal (primers designate sequence to be copied)
Elongation – Sample is heated to the optimal temperature for a heat-tolerant polymerase (Taq) to function (~75ºC)
What enzyme is used in PCR and why?
Taq polymerase is an enzyme isolated from the thermophilic bacterium Thermus aquaticus
As this enzyme’s optimal temperature is ~75ºC, it is able to function at the high temperatures used in PCR without denaturing
How does Taq polymerase work?
Taq polymerase extends the nucleotide chain from the primers – therefore primers are used to select the sequence to be copied
What is transcription?
Transcription is the process by which an RNA sequence is produced from a DNA template
- What is the role of RNA polymerase in transcription?
RNA polymerase separates the DNA strands and synthesises a complementary RNA copy from one of the DNA strands
- What happens once the strands are separated in transcription?
When the DNA strands are separated, ribonucleoside triphosphates align opposite their exposed complementary base partner
- How does RNA polymerase create the mRNA copy during transcription?
RNA polymerase removes the additional phosphate groups and uses the energy from this cleavage to covalently join the nucleotide to the growing sequence
- What happens once transcription is complete?
Once the RNA sequence has been synthesised, RNA polymerase detaches from the DNA molecule and the double helix reforms
What is a gene? (referring to transcription?
The sequence of DNA that is transcribed into RNA is called a gene
What is the strand that is transcribed called?
The strand that is transcribed is called the antisense strand/template strand and is complementary to the RNA sequence
What is the strand that is not transcribed called?
The strand that is not transcribed is called the sense strand and is identical to the RNA sequence (with T instead of U)
Where does transcription occur?
Transcription of genes occur in the nucleus (where DNA is), before the RNA moves to the cytoplasm (for translation)
What does the base sequence of an mRNA molecule encode?
The base sequence of an mRNA molecule encodes the production of a polypeptide
What are codons, what is their purpose?
The mRNA sequence is read by the ribosome in triplets of bases called codons
What does each codon code for?
Each codon codes for one amino acid with a polypeptide chain
What does the order of codons determine?
The order of the codons in an mRNA sequence determines the order of amino acids in a polypeptide chain
What is the genetic code?
The genetic code is the set of rules by which information encoded within mRNA sequences is converted into amino acid sequences (polypeptides) by living cells
What does the genetic code identify?
The genetic code identifies the corresponding amino acid for each codon combination
How many codon possibilities are there and why?
As there are four possible bases in a nucleotide sequence, and three bases per codon, there are 64 codon possibilities (43)
What does the coding region of an mRNA sequence always begin and end with?
The coding region of an mRNA sequence always begins with a START codon (AUG) and terminates with a STOP codon
What does the genetic code typically show?
Typically the genetic code shows the codon combinations expressed on an mRNA molecule
How does the mRNA copy differ from the coding strand?
These sequences are identical to the mRNA codons with the exception of thymine (T) being present instead of uracil (U)
Why is DNA termed a semi-conservative process?
DNA replication is a semi-conservative process, because when a new double-stranded DNA molecule is formed:
One strand will be from the original template molecule
One strand will be newly synthesised
Why is the newly formed DNA semiconservative?
This occurs because each nitrogenous base can only pair with its complementary partner
Adenine (A) pairs with thymine (T)
Cytosine (C) pairs with guanine (G)
What will the two newly formed DNA strands be like?
Each new strand formed will be identical to the original strand separated from the template
The two semi-conservative molecules formed will have an identical base sequence to the original molecule
What experiment confirmed the semi-conservative nature of DNA?
The theory that DNA replication was semi-conservative was confirmed by the Meselson-Stahl experiment in 1958
What 3 hypotheses were proposed before meselson and stahl?
conservative, semi-conservative and dispersive mdel
What does the conservative model propose?
An entirely new molecule is synthesised from a DNA template (which remains unaltered)
What does the semi-conservative model propose?
Each new molecule consists of one newly synthesised strand and one template strand
What does the dispersive model propose?
New molecules are made of segments of new and old DNA
What did M and S use to test their theory?
Meselson and Stahl were able to experimentally test the validity of these three models using radioactive isotopes of nitrogen
Why did M and S use nitrogen?
Nitrogen is a key component of DNA and can exist as a heavier 15N or a lighter 14N
- What was the first step of the process? M & S
DNA molecules were prepared using the heavier 15N and then induced to replicate in the presence of the lighter 14N
- How were the DNA samples then separated and why? M & S
DNA samples were then separated via centrifugation to determine the composition of DNA in the replicated molecules
What were the results of M and S after one division?
After one division, DNA molecules were found to contain a mix of 15N and 14N, disproving the conservative model
What were the results of M and S after two divisions?
After two divisions, some molecules of DNA were found to consist solely of 14N, disproving the dispersive model
What is translation?
Translation is the process of protein synthesis in which the genetic information encoded in mRNA is translated into a sequence of amino acids on a polypeptide chain
- What is the first step of translation?
Ribosomes bind to mRNA in the cytoplasm and move along the molecule in a 5’ – 3’ direction until it reaches a start codon (AUG)
- What happens once the mRNA is attached?
Anticodons on tRNA molecules align opposite appropriate codons according to complementary base pairing (e.g. AUG = UAC)
- What is the role of tRNA?
Each tRNA molecule carries a specific amino acid (according to the genetic code)
- What is the role of ribosomes?
Ribosomes catalyse the formation of peptide bonds between adjacent amino acids (via condensation reactions)
- Does the ribosome move or the mRNA strand? When does the process stop?
The ribosome moves along the mRNA molecule synthesising a polypeptide chain until it reaches a stop codon
- What happens once the stop codon is reached?
At this point translation ceases and the polypeptide chain is released
What are the key components of translation?
Messenger RNA (goes to…) Ribosome (reads sequence in …) Codons (recognised by …) Anticodons (found on …) Transfer RNA (which carries …) Amino acids (which join via …) Peptide bonds (to form …) Polypeptides
Mnemonic: Mr Cat App
What is a special property of the genetic code?
The genetic code is universal – almost every living organism uses the same code (there are a few rare and minor exceptions)
Due to the genetic code being universal, what can be done between species?
As the same codons code for the same amino acids in all living things, genetic information is transferrable between species
Why is transferring genes between species useful?
The ability to transfer genes between species has been utilised to produce human insulin in bacteria (for mass production)
- Where do scientists get the insulin-producing cell?
The gene responsible for insulin production is extracted from a human cell
- What is done to the insulin-producing cell?
It is spliced into a plasmid vector (for autonomous replication and expression) before being inserted into a bacterial cell
- What is then cultured?
The transgenic bacteria (typically E. coli) are then selected and cultured in a fermentation tank (to increase bacterial numbers)
- What is the final result of insulin production?
The bacteria now produce human insulin, which is harvested, purified and packaged for human use (i.e. by diabetics)
What is mRNA?
mRNA is a complementary copy of a DNA segment (gene) and consequently can be used to deduce the gene sequenc
What is the mRNA transcript organised into?
The mRNA transcript is organised into triplets of bases called codons, and as such three different reading frames exists
What does the mRNA transcript start with?
An open reading frame starts with AUG and will continue in triplets to a termination codon