Sunday 10/04/15

exploring tSNE (11 am -3:50 pm)

  • looking at L7 data
  • see the expected correlations (here in 3D)
  • can also see dataset biases (these go away on z-normalization).
  • 1000 gene data
  • not so good for chromatin structure yet
    • algorithm cares about absolute orientation (forward and backward slanty 1s are different, 6s and 9s are different).
    • could do some sort of global rotation to fix this, which would rotate all 1s in same direction
    • still wouldn’t capture features likes lots of protrusions — its hierarchical clustering of absolute distances of the measurements.
  • obviously our data descriptors like volume still have interesting affects on the data
Posted in Summaries | Comments Off on Sunday 10/04/15

Thursday 09/24/15

1:00 pm – 7:00 pm, 8:30 pm – 9:45 pm

job apps

  • revising research statement (remotely 9:00 am – 1:00 pm)
  • sent to a few colleagues and XZ for comments
  • working on teaching statement 5:00 pm – 7:00 pm


  • BB testing KD
  • 70% KD efficiency (down from 80-90)
  • options
    • have more of the prepped sample from last time, can run as a technical replicate at higher concentration to address the 0 reads issue.
    • might have more RNA in the freezer (not sure about this)
    • have more cDNA from QPCR experiments. should be able to prep this for sequencing. won’t have the easy length control. Maybe not a big problem for the reads — might have to do some adjustment of PCR amplification step.
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Protected: literature notes

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Wednesday 09/23/15

9:00 am

job apps

  • working on figures for research proposal, 9:00 am


  • working on revisions.
Posted in Summaries | Comments Off on Wednesday 09/23/15

Tuesday 09/22/15

9:00 am

Job Apps

  • working on figures for Research Statement (11a – 2p, 8p-10p)
  • some progress on corresponding text.


  • tweaking alternative version of EDF 3. (9:30 – 10:45am)
  • revising response to reviewer letter 2pm – 4:30 pm, 6:00pm – 7:30pm

STORM hal updates

  • brief tutorial from Hazen (11:15 – 11:30)
  • see hb-storm-control (development branch) on STORM2
  • new settings (examples in Hazen documents)
  • camera has multiple feeds
    • settings file can specify feeds
    • these can display just a sub-field of view
    • these can also integrate a given number of frames
    • or just display every Nth frame
    • they can be told to save, they save with the name of the main file plus appending a user defined flag which you name the feed
    • camera1 (the whole field) is also saved by default unless you set save to False.
  • steve settings are now all apart of hal settings
  • no longer will have separate branches for separate setups, everything is in the parameters file
  • Hal needs to be running first (or right click steve and select Query Hal) to get the objective and settings
  • still need to add the 1.5x slider option to new hal parameters (from my Steve parameters).
  • Hal/steve/dave all reset the python paths so you can have multiple functional copies of storm-control
  • movie saving is now handled by main thread, not by camera. Keep an eye out for movies that drop frames or hang while saving (if camera drops frame hal might not know).
Posted in Summaries | Comments Off on Tuesday 09/22/15

Monday 09/21/15

9:00 am – 7:00 pm, 9:00 pm – 10:00 pm

Manuscript revisions

  • revising main text in response to XZ comments
  • figure adjustments in response to XZ comments
  • discussed main text revisions
    • simplified Ph KD discussion
    • simplified PRE discussion
    • simplified fixation controls and discussion
  • further formatting of main text
  • adjusted examples in fixation controls, show more detail.
  • sent to collaborators
Posted in Summaries | Comments Off on Monday 09/21/15

Sunday 09/20/15

10:30 am – 7:00 pm

Job apps

  • writing research statement

Figure revisions for chromatin

  • see XZ notes


  • DNA digestion following IVT (dilute 2x add DNase I from NEB IVT quick kit)
  • RNA purification
  • quantify yield
  • transfect new KC cells. (5:15 pm)
Posted in Summaries | Comments Off on Sunday 09/20/15

Saturday 09/19/15

10:40 am – 1:30 pm

New Ph KD

  • set up new Ph KD with last drops of previous dsRNA batch for ph-p and ph-d (well, I still have some ph-d left). Also did Pc KD as a bonus.
  • ran 10% short on ph-p. And I’m not sure if this dsRNA is still up to snuff, its been thawed at least 5 times and in the -80 for over a month.
  • knockdown started at 12:10 pm.
  • started prepping new dsRNA for ph-p and ph-d.
    • PCR / cleanup column doesn’t seem to be ideal — I spec’d the DNA (~70ng/uL and 160 ng/uL respectively for ph-p and ph-d) and the 230 peak is as high as the 280.
    • set up T7 reactions anyway. Might run a gel after the DNase treatment and RNA cleanup (do tomorrow?)
  • should probably do another round of QPCR on these RNA isolations before deep sequencing to check KD given issues.

Job app stuff

  • working on revised statement.
  • updated CV with CB comments.
Posted in Summaries | Comments Off on Saturday 09/19/15

Friday 09/18/15

9:05 am – 5:15 pm

Chromatin manuscript

  • revising ED fig 3
    • I think we can make the fig 1 images work
    • might need to do denoising requested by reviewer
    • also tuned up previous examples and improved contrast (still only 1 example per each, was requested 3).
  • finished revised letter fig 2 with 3 3×3 panels comparing centromeric, repressed and active chromatin
  • added 300 cells into ED fig 4 and reformatted the fonts and labels
    • modified script to output “A-” and “I-” to meet the very latest of the naming conventions (didn’t propagate into allData structure yet).
  • revised main text, discussed changes with BB. Sent to XZ
  • XZ requests more deep sequencing.
Posted in Summaries | Comments Off on Friday 09/18/15

Thursday 09/17/15

9:15 am – 10:00 pm


  • revising main text

chromatin experiments

  • helping BB do deep-seq prep
  • KapaQuant results:
  • concentrations lower than expected, (only most concentrated of the dilutions look good)
  • suspect a problem with the beads
  • Tapestation quantification — no detectable library smear. Prep failed.
Posted in Summaries | Comments Off on Thursday 09/17/15