extra bits - mutations, PCR, operons

SBI4U - Howes

large scale vs point mutations

Large Scale: involving whole genes or regions of a chromosome

Point: Involving single nucleotides

3 causes of a point mutation

• substitution of one base pair for another

• insertion or deletion of a single base pair

• inversion of 2 adjoining base pairs

kinds of point mutations (4)

1. Silent Mutation: no change in polypeptide

2. Missense Mutation: same codon frame, different amino acid created

3. Nonsense Mutation: results in a premature stop codon (not an animo acid)

4. Frameshift Mutation (insertion/deletion): shifts the entire reading frame over by one — every downstream a.acid is altered

Mutation consequence in somatic vs germ cells (gametes)

Somatic: if recognized= apoptosis, if not recognized = uncontrolled growth and cancer

Gamete: All of the offspring’s cells will carry the mutation

Causes of mutations

Spontaneous: error in DNA replication (DNAP III)

Induced: external factors that can impact DNA

PCR stands for:

Polymerase Chain Reaction

What is PCR?

A technique to make many copies of a specific DNA region in vitro

PCR, like DNA replication, still requires:

a DNA polymerase enzyme

The DNA polymerase used in PCR is called ______________, and is special because:

Taq polymerase: it's heat-stable, so doesn't denature in the high temperature used in PCR

why is high-temperature repeatedly used in PCR?

to denature the template DNA, separating its strands

Taq polymerase still needs a _______ to start synthesizing. PCR primers are ______ pieces of ____

PRIMER, short, DNA

How does the experimenter choose the region of DNA to be copied?

By using two primers that flank the target region — the primers are given sequences that will bind to complementary base pairs on the template DNA

3 steps of PCR

1. Denaturation (96°C): separate DNA strands

2. Annealing (55-65°C): allow primers to bind

3. Extension (72°C) taq polymerase extends primers

explain the behaviour of the trp operon in low and high lvls of tryptophan

HIGH: tryptophan binds to the repressor, activating it. Once activated, the repressor binds to the operator, blocking transcription

LOW: tryptophan isn't there to bind to the repressor, so transcription continues and results in the synthesis of tryptophan

explain the behaviour of the lac operon in low and high lvls of lactose

HIGH: lactose binds to the lac repressor, which deactivates it. Transcription continues and results in the synthesis of lactose-metabolizing enzymes.

LOW: without lactose, the lac repressor is free to bind to the operator, blocking transcription

Why does the repressor binding to the operator stop transcription?

It blocks the RNA polymerase from binding!