D1 - SL

What is DNA replication?

Production of new strands of DNA with identical base sequences to existing strands

What does it mean that DNA replication is 'semi-conservative'?

Each replication results in two double-stranded DNA helices, with each containing one strand from the original DNA and one newly made strand

How does complementary base pairing facilitated the semi-conservative nature of DNA replication?

Each newly separated single strand provides the template for the opposite strand

What is the role of the helicase enzyme in DNA replication?

Unwinds and breaks hydrogen bonds between DNA strands

What is the role of the enzyme DNA polymerase in DNA replication?

Attaches nucleotides to the growing complementary strand, using the existing template strand as a guide. Proofreads and removes non-complementary nucleotides.

What is the purpose of the polymerase chain reaction (PCR)?

Amplify the amount of DNA via artificial DNA replication in a laboratory setting

What are the five ingredients needed for a PCR reaction?

Small amount of DNA

Taq DNA polymerase

Thermocycler

Free DNA nucleotides

'Target specific' primers

What are the three steps of PCR, including the target temperature?

Denaturation @ 95℃ - heat breaks the hydrogen bonds between nucleotides, separating DNA helix into two single strands.

Annealing @ 54℃ - primers bind to the target sequences of separated strands

Extension @ 72℃ (optimum temperature for Taq polymerase) - Taq polymerase synthesises new DNA strands by adding complementary free nucleotides

REPEAT (25-30 rounds)

What is an example of the application of PCR?

Covid testing i.e. exponentially increase small amounts of Covid RNA so the virus can be detected, even in low levels

What is the goal of gel electrophoresis?

Separate DNA fragments based on their size to visualise an individual's unique DNA banding pattern

What are the seven steps of gel electrophoresis?

1. Amplify the desired DNA via PCR

2. Cut DNA into short fragments with restriction enzymes

3. Create an agarose gel with small pores and wells

4. Insert DNA into wells with dyes

5. Run an electrical current. DNA has a negative charge and will be attracted to the positive electrode. Separates DNA fragments by size (smaller fragments travel further)

6. Stop current before the smallest strand reaches the end

7. Use a stain or fluoresence to visualise

What are some applications of gel electrophoresis?

Paternity testing, DNA profiling

What is transcription?

Synthesis of a strand of messenger RNA from template DNA for a single gene.

What are the four functions of RNA polymerase in the process of transcription?

- unwind DNA helix into two strands

- move along template strand positioning complementary RNA nucleotides

- create phosphodiester bonds between RNA nucleotides

- detach RNA from DNA template so DNA can reform a helix.

What are the complementary base pairing rules in transcription?

DNA adenine -> RNA uracil

DNA guanine -> RNA cytosine

DNA cytosine -> RNA guanine

DNA thymine -> RNA adenine

What is the relationship between gene expression and protein synthesis?

Protein synthesis is the result of gene expression.

e.g. the transcription of one gene of DNA into mRNA allows a polypeptide to be created by translation. After modification, this becomes a protein

What is the difference between sense and anti-sense strands in transcription?

Sense (AKA coding) strand = sequence matches the RNA transcript (except that RNA has U instead of T), not used as a template by RNA polymerase.

Anti-sense (non-coding, template) strand = template for RNA polymerase during transcription, has complementary bases to the mRNA

What is translation?

The synthesis of polypeptides from mRNA at ribosomes i.e. base sequence of mRNA is translate into the amino acid sequence of a polypeptide

What are the three main types of RNA?

Messenger RNA

Ribosomal RNA

Transfer RNA

What is a codon?

Every three bases of mRNA. Each codon codes for one amino acid.

What is an anti-codon?

A trinucleotide sequence at the end of tRNA that is complementary to a relevant mRNA codon. Helps the tRNA find the correct mRNA codon.

What is the role of complementary base pairing in translation?

Complementary base pairing between mRNA codons and tRNA anti-codons allows the correct amino acid to be delivered to the correct codon.

What does it mean to say that the genetic code is degenerate and universal?

Universal = same codons code for the same amino acids in every living organism

Degenerate = more than one codon codes for the same amino acid e.g. 64 codons but only 20 distinct amino acids.

What is the function of tRNA in translation?

Delivers the correct amino acid to the codon on the ribosome.

What are the three sites of the ribosome?

Arrival site

Peptidyl site

Exit site

What are the six main events in the repeating steps of translation?

1. tRNa carrying an amino acid arrives at the A (arrival) site.

2. This amino acids forms a peptide bond to connect it to the elongating polypeptide chain

3. tRNA is shifted to the P (peptidyl) site once they are 'holding' the chain, as the ribosome moves down the mRNA (stepwise movement)

4. Once another tRNA arrives at the A (arrival) site and a new peptide bond forms, the polypetide chain will transfer to this new tRNA

5. At that time, the original tRNA is shifted again by the ribosomal movement into the E (exit) sit

6. The tRNA will leave the ribosome

What direction is the mRNA read by the two ribosomal subunits?

5' to 3'

What part of the ribosome does the mRNA bind to?

The small subunit of the ribosome

How many tRNAs can bind simultaneously to the large subunit of the ribosome during translation?

2

What is a mutation?

Random and rare change to genetic material at the molecular level

What is a point mutuation?

A form of mutation where only one single nucleotide is impacted

What is a single-nucleotide polymorphism?

A mutation that changes one nucleotide and creates a new allele for that gene

What are the three types of gene mutations?

Substitutions

Insertions

Deletions

What are the three broad categories of mutations?

Chromosomal - a large section of DNA is damaged

Gene - a small section of DNA within one single gene is altered

Non-coding - a small section of DNA that does not code for proteins is altered

What is a substitution mutation?

a mutation that exchanges one nucleotide base for another

What are the three possible outcomes of a substitution point mutation?

Missense

Nonsense

Silent

What is a missense substitution point mutation?

When the one nucleotide base change leads to a different amino acid being expressed, but all other amino acids are unchanged. The protein is altered yet may be somewhat functional depending on R group similarity.

What is a nonsense substitution point mutation?

When the one nucleotide base change leads to a stop amino acid being expressed. All amino acids after this mutation are lost, leading to a non-functional protein that is broken down.

What is a silent substituion mutation?

When the one nucleotide base change leads to a same amino acid being expressed, due to the degeneracy of the genetic code. Protein remains unchanged and is fully functional

What is an insertion mutation?

A mutation that adds one or more based to the gene, often resulting in a frameshift that changes every codon after the mutation

What is a deletion mutation?

A mutation that removes one or more nuclotide bases from the gene, often resulting in a frameshift that changes every codon after the mutation

What do insertion and deletion mutations usually result in?

A frameshift. All codons after the mutations are changed, leading to the expression of different amino acids and usually a non-functional protein.

Why does a single point mutation result in sickle cell anemia?

A single substitution of a Thymine base to an Adenine base in the gene for Hemoglobin B subunit expresses a different amino acid, leading to an altered shape of haemoglobin and causes a clustering of proteins.

What mutation is Huntington's Disease a consequence of?

An insertion mutation. The repeated insertion of trinucleotide repeat (CAG) into the gene HTT causes Huntington's disease, after 40+ repeats.

What mutation is the delta 32 mutation of the CCR5 gene an example of?

Deletion. This means that CCR5 receptor (which HIV can bind to) on helper T cells (WBCs) will not be expressed -> HIV resistance.

What is a mutagen?

Any substance that causes mutations

What is satellite DNA?

Non-coding DNA that plays a structural role by forming the centromere during cell division?

What are some examples of chemical mutagens?

Carnicogens e.g. cigareete smoke

Industrial benzene, found in acetone

What are some examples of mutagenic forms of radiation?

UV, x-ray, nuclear power plants

What is an internal cause of mutation?

An error during DNA replication or repair (e.g. meiosis or mitosis) causes the wrong base to be placed in -> results in a substitution point mutation.

What does it mean to say mutations are random?

They can occur anywhere in the genome, but some areas of DNA are more prone e.g. uncoiled DNA

How do mutations affect somatic cells?

Will impact that person but will not be passed onto offspring. Often develops into a cluster of abnormal body cells (tumour) but will impact individuals, not the populations.

How do mutations affect germ cells?

If mutations occur during meiosis or in existing sex cells, the original DNA that makes up the zygote will be altered and passed onto all cells of the offspring.

What role does mutation play in natural selection?

Mutation creates new alleles of genes and is therefore the original source of all genetic variation. Although most mutations are either harmful or neutral for an individual organism, in a species they are essential for evolution by natural selection in the long-term