DNA Unzipping

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DNA Unzipping by Mind Map: DNA Unzipping

1. Tethering Conditions

1.1. Milestone to shoot for

1.1.1. With dsDNA (4.4 or 1.1 kb): good results almost every time

1.1.1.1. Good tether: stuck ratio

1.1.1.2. Good over all tether density (a couple / field of view)

1.2. Necessary footholds

1.2.1. Good monodisperse beads

1.2.1.1. Dilution technique

1.2.1.2. Sonication conditions

1.2.2. Good sample chambers

1.2.2.1. cleaning glass necessary

1.2.2.1.1. sonicate in alconox and water for 3min and rinsing numerous times. Blow dry.

1.2.3. Good blocker

1.2.3.1. prevents stuck beads

1.2.3.2. still permits tethering

1.2.4. Good antibody

1.2.4.1. diluted with PBS

1.2.4.1.1. tethers possible

2. SDM Code

2.1. Probably will require overhaul, once we start using it again

3. Shotgun DNA Mapping (specific aim 1)

3.1. Necessary foundation

3.1.1. DNA Constructs

3.1.1.1. Plasmid only construct

3.1.1.1.1. Success via gel observation

3.1.1.1.2. success via tethering?

3.1.1.1.3. not unzipped yet

3.1.1.2. shotgun clones construct

3.1.1.2.1. Success via gel observation?

3.1.1.2.2. Not necessary for biophys poster. Necessary for CAREER and other grants

3.1.2. Optical Tweezers

3.1.2.1. Hardware

3.1.2.1.1. Necessary tasks

3.1.2.1.2. Things to probably purchase

3.1.2.2. Software

3.1.2.2.1. OT Control code

3.1.2.2.2. Data analysis code (converting raw to force versus unzipping index)

4. Pol II unzipping (specific aim 2)

5. Unnatural DNA unzipping (benner discussion)

6. Telomere Unzipping