Lecture 1 - Overstretched DNA

Question 1

Describe Optical Tweezers

Answer 1

-Highly focused Laser Beams that can trap particles at their focal point

-Particle/bead coated in streptavidin, DNA ends coated in biotin and tethered together. (Held together by biotin-streptavidin bond)

-DNA can be stretched by moving laser beams, using mirrors.

Question 2

Define Contour Length and Persistence length.

Answer 2

Contour Length (Lc): The maximum end-to-end length without deformation

Persistence Length (Lp): The length over which the molecular axis of an object approximates as a straight line

Question 3

What is overstretching and at what point does it take place?

Answer 3

Overstretching is when DNA elongates without any additional force added. This happens at a force of 60 pN

Question 4

What are the 3 models to explain overstretching?

Answer 4

Stand Peeling
Melting bubbles
S-DNA

Question 5

What does experimental data show about the models?

Answer 5

At end open DNA: Strand peeling
At end closed DNA: Melting bubbles

At high ionic concentrations or GC-rich areas: S-DNA

Question 6

What is TFAM and what does it do?

Answer 6

Mitochondrial Transcription factor A compacts mitochondrial DNA by creating hinges in the DNA, making it more flexible.

It also rapidly slides across DNA (10e-5 nm2/s), regulating transcription (more sliding, less transcription)

Question 7

What is the role of torsional stress?

Answer 7

A negative change in writhe (negative supercoiling) results in a higher Lc, lower Lp, increasing flexibility.

Negative supercoiling reduces TFAM sliding, promoting transcription.