The Joy of Koching (080613)

8/14/2019 The Joy of Koching (080613)

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The American ScienceClassic

Revised and Expanded

With Over 4 500

THE ALL-PURPOSE

OFKOCHING

by ANTHONYSALVAGNO


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Let us give thanks

Kochlab Nation Steve Koch (founder

and godfather) Larry Herskowitz

Patrick Jurney Linh N Le Diego Ramallo-Pardo Nas Manole Caleb Morse

Adam Wise (NSMS)

The VIP List Osley Lab Lopez Lab Nick Carroll

Chris Lorenz Richard Yeh Karen Adelman

The Beautiful People


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Bread Making 101


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Extracting Chromatin from livingyeast cells

Tethering extracted Chromatin toslide

Overview ofPart Part

Korber et al. MCB . (2004). 24: 10965-

Preheat the Oven to


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1st knead the dough

Need Pho5 plasmid Contains well documented nucleosome

locations

CellW

Mock yeast

Nucle To digest chromatinone needs accessto nucleus

1st attemptsinvolved creation ofspheroplasts

Current attempts


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next allow dough to rise

After extraction, tetheringcommences Chromatin needs to be prepared in a

tethering construct (dig labeling on oneend and biotin labeling on the other).

Above: Tethering Constuct(Steve Koch)

Left: Side profile of tetheredparticles ready for


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and bake for 30-40 min

Optical Tweezers

Allow manipulationof DNA tethers

Microfluidics

Allow automation oftethering process

Provides backboneto many futureexperiments

The Tools of the

Tweezing courtesy of Microchannel


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Make sure to check

Building OT Need laser

Beam steering

Data Acquisition Microscope

Simple Design by Block and


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Place on wire rack to cool.

Building Microchannels Double Stick Tape Method PDMS 1.0

Using Photoresist

Greatly improves precision of channel making Can create all types of different shapes and sizes

for use in various experiments

Mask Design by Patrick


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Presentation is important

Simulations:

Thermodynamic calculations of nakedDNA unzipping

Monte Carlo simulations of RNAPolymeraseTranscription on chromatin

Molecular Dynamics of protein-DNAinteractions Histone-DNA and RNA Pol II-DNA

Atomistic MD for microsecond timescale


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More Recipes

Shotgun ChromatinMapping

Create shotgun

chromatin fragmentsusing I-PpoI

Tether fragments

Unzip fragments


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and even more!

$6 Million (Man) Tweezers Beam Steering

AOD Mirror Galvanometer

Better Data Acquisition Feedback

Micro- (and nano-) fluidic experiments PDMS stamping Flow experiments

Simulations Combination of atomistic and CG MD


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The End

Documents

The Joy of Koching (080613)