9:30 am -5:50 pm, 7:30 pm – 12:00 am
- Now black behaves properly but yellow and blue are giving issues
- yellow not equilibrating?
- not sure what’s going on with blue, will require further investigation. Possible that large domains are swallowing smaller ones and growing too fast.
- also possible that large domains are just splitting into multi-rosettes and small domains stay as singles.
- ‘WholeNucVOp5′ – 10% sticky blue, blue attraction 3.0 repulsion 3.0
- ‘WholeNucVOp6′ – yellow back to no stiffness, run for 200 steps.
- this does (as before) a good yellow black split.
- New approach — successively add more sticky sites to BLUE. see if we get a better saturated state if it folds in steps.
Discussion with Xiaowei
- XZ doesn’t like the yellow model as a faster equilibration to Eq. Globule from Fractal globule (due to energy or chain-crossing). Too much of a polymer explanation, not enough useful biology.
- probably right, I think the modeling is actually rewarding and complex enough just with the blue.
Multiplex single-molecule interaction profiling of DNA-barcoded proteins
Nature 2014, Church lab, Gu et al
presented by Pallav
- current approaches = protein vs. library
- rather do an everyobdy to everybody screen. One pot analysis not well-based screen
- single-molecular interaction sequencing (SMI-seq)
- barcode proteins with DNA.
- on the mRNA add a barcode and a stalling tag downstream of a polypeptide linker. The ribosome now combines the mRNA and the protein together. (during an in vitro translation reaction following an in vitro transcription reaction).
- can now mix proteins
- barcoded RNA annealed to RNA with little tags.
- create dilute array of polonies in acrylomide gel. Can now sequence.
- have two different pools with two different primers. Only the ones with one of each primer can be decoded.
- challenges: very different yields of different complexes. Can count total spots to get estimate of concentrations and thus get estimates of binding efficiency ?
ligand binding screening
- small molecule binding to GPCR (g-protein coupled receptor) measure affinity based on GPCR’s change in affinity for binding protein arrestin. different ligand in each well.
library to library screening
- library 1 antibody domains (a bunch of mutated variable forms)
- library 2 human proteins
- test 200 x 60. With Illumina reading propose could test 10,000 x 10,000
10:00 am – 2:30 pm, 4:00 pm – 11:00 pm
- lab meeting (see notes)
- journal club (see notes)
- discussion with XZ
10:30 am – 6:20 pm, 10:30 pm – 12:00 am
- Anaconda called from sbatch actually worked to get save data.
issues with current simulations
- black chromatin starts at .4 (and stays there).
- writing new template to do the the fractal globule 2 step formation first before starting simulation
- this is actually relatively slow, we may not want to do it fresh each time.
Summary of recent simulations
- WholeNucV6p15 – 30+ runs of basic blue/black/yellow
- yellow and black split off a bit too late, but with appropriate difference in slope
- both yellow and black a bit too relaxed — yellow .5 black .4
- blue is holding at correct Rg slope of .2 and a reasonable density, looks
- running on Odyssey rpt of V6p15 with stiffer yellow and slightly fewer sticky blue
- testing on Monet new pipeline to generate and relax fractal globule all in same script (V7)
- V7p2 looking a bit better:
Running V8p1 — trying to get black to initialize closer to .3 than .4. I think this is because the Random Walk Polymer initiation starts entangled. Trying a simulation with a condensed regular coil.
- discuss PH project follow up
10:00 am – 1:00 pm, 7:00pm – 9:00pm,
- troubleshooting running polymer simulations on Odyssey GPU
- issues using polymer load and save commands — requires joblib
- testing out with Anaconda via sbatch command