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2020-04: 2 entries 2020-05: 2 entries 2020-06: 4 entries 2020-07: 9 entries 2020-08: 19 entries 2020-09: 7 entries 2020-10: 6 entries

2020

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April

Error: Kernel died

When running my batch for running PharaGlow, I got the following message for the recording “EB0043c”

Autosave too slow: 8.01 sec, over 25% limit. Backing off to 60 sec
Kernel died while waiting for execute reply.

Out of memory or reach the time limit of running a kernel(~8 hours) ?

https://www.kaggle.com/c/data-science-bowl-2019/discussion/126052

A too old version of anaconda / jupyter notebook ? https://stackoverflow.com/questions/49326164/jupyter-notebook-dead-kernel

2020/10/30 09:23 · ebonnard

Staging INF18 worms

Aim

How many hours after egg laying could we obtain young adults INF18 worms ?

Protocol

See how to stage worms in wiki protocols

20201027: Synchronization (3x ~10 worms per plate)

  • Worms from healthy maintenance plate
  • Pick young healthy gravid adults worms onto intermediate plates
    • 2x10cm-diameter plates
    • 30 worms per plate
    • Let worms go around for 60 minutes
    • Observation: 60Min. is too long. Worms make new eggs that may stick to them when I pick them. ToDo: try 30-40 Min.
  • Transfer worms onto the syncing plates
    • 6x 6cm-diameter plates
    • (1) (2) (3) worms per plate
  • Egg laying from (1) 17:15 - (6) -18:50 = ~1h20
  • Incubate at 20C

Results

plate 1 Date HH:MM cum. time Observations
Start Egg laying 27.10.2020 17:30 -1h20 x healthy gravid worms
End Egg laying 27.10.2020 18:50 0 many eggs
28.10.2020 09:00 14 small worms with green pharynx
28.10.2020 21:00 26h
29.10.2020 09:00 38h
L4 30.10.2020 09:00 62h moon shape visible
Young adults 30.10.2020 14:15 67h no more l4-moon, scarce eggs

idem for plate 2 (17:34-18:51) and plate 3 (17:36 - 18:46)

Conclusion

  • Inf18 worms need ~67hours to become young adults
2020/10/27 02:37 · ebonnard

One-on-One Meeting (26.10.2020)

From the recordings on the 13.08.2020 (EB0023-26) and 20.08.2020 (EB0031-38), we learnt that worms responses to vibration by 1/ increasing their reversal rate, 2/ increasing their instantaneous velocity (EB0034, S1: ~x8) and 3/ decreasing the pumping rate (EB0034, S1 and S2: ~50%). Next experiments with “cleaner” conditions aimed to replicate these results. However the pumping response were much lower (E.g. EB0041-42). Since I suspected a variation in the transmission of the vibration from the piezo buzzer to the plate, I have integrated in the set up a piezoelectric sensor to assess this transmission ( EB0043-44). As, I saw a high variation in intensity (but not frequency), I plan to work on a stable version of the plate adapter.

  1. Protocol and set-up development for the behavioral experiments to gain reproducibility
    • Parameters selected - where worms respond to vibration by decreasing their pumping rate, moving faster and increasing their probability of reversal
      • stimulus: vibration at 630Hz, 20Vpp, 1s duration, 30s to 3Min. ISI, x15-x5
      • in 6cm plate: 70uL lawn, ~100 worms
      • See more parameters and how they have been adjusted during the last experiments in the reports in wiki
    • Stimuli triggering automated coupled to the camera.
    • Piezoelectric sensor added to assess vibration transmission from the piezo buzzer to the plate
      • With the current plate adapter, the transmission may change sensibly between recordings and possibly during a same recording.
      • ToDo: The adapter must be improved and tested by the method recently developed.
        • Alternative post recording, with analysis:
          • ToDo: check if there is no “miss pump” in PharaGlow: compare pumping rate from visual pumping and from PharaGlow
          • Get statistics
          • Sub-groups divided according the velocity response: 1/ increase –> pumping rate 2/ no change –> control
  2. Mutant strains selected, requested,(partly) received and/or in “preparation” for next behavioral and GCaMP imaging experiments
  • INF18 = priority
    • Behavior experiment for both inf18 and gru101 worms in the same day
  • For GCaMP imaging: [[wiki:documentation:strains|swf62]] (NSM)
    • ToDo = plan with Luis a training week to image calcium signal in single freely moving worm while stimulation. Use joystick to manipulate the stage
  • Jun is crossing the strains. Should I dedicate time now to learn worms crossing with him? = Yes
  1. Analysis:
    • A logger added to PharaGlow (ToDo: add to Github) and my analysis script to keep track errors (by waiting to solve them with you (-: )
      • ToDo: send error messages from PharaGlow to Monika
      • ToDo: change PharaGlow parameters “memory” and lower the search range. Minimal length should be higher than the memory. If memory too smal, small dashed line in the trajectories
        • Memory was equal to 200, for EB0045-48 try 100 (Monika: 30 / 100)
        • Search range = 60, try 30 (Monika: 10 / 60)
    • Reversal, pumping rate and velocity are monitored: events plot and peristimuli data including normalisation > Need more work to integrate all together.
      • Get statistics: calculate the error of the mean, T-Test, number worms as function of time
      • Get info from control: normalize the control as I did for the peristimulus data but with “false” stimulus
    • Automation of the analysis via batch scripts for PharaGlow and my analysis. > Got recent errors “kernel dead” to solve
      • This error could come from an out of memory, with Monika, move the analysis on axon
  2. Application to the IMPRS (deadline 15 Nov.), following by interview training and poster presentation. More info here. > The online application requires more work, to do first (?)
    1. Contact Martha Daniel to help each other with the interview
2020/10/25 10:49 · ebonnard

mec-10 and mec-4 mutants

2020/09/16 07:02 · ebonnard

Pumping Analysis (EB0040-41)

The stimulus parameters have been validated by the previous experiments (plate adapter, frequency = 630Hz, duration =1s, intensity= 20Vpp ; ISI=1Min or 3Min instead of 30s), as well as the imaging conditions to detect pumping events (Zeiss “32x”, 30Hz, V-OP50= 60uL instead of 40uL). The ISI is longer in order to see when the pumping rate return to its baseline and the Op50 seeding volume is higher to bit a little bit larger than the field of view of the camera and therefore avoiding interface. In addition, the control consists now of the 3 first minutes of recording before the stimulation.

EB0039: ISI = 3Min.

20200914_eb0039_pumping.pdf

EB0040: ISI = 1Min.

20200914_eb0040_pumping.pdf

How long did acclimation time lasted from the previous experiments ? = for next experiment, let the worms acclimate for 2h

acclimation
EB0031 1h45
EB0032 1h45
EB0033 2h40
EB0034 3h20
EB0039 1h15
EB0040 1h26

See Experiment Log

2020/09/15 01:13 · ebonnard
wiki/documentation/elsa.txt · Last modified: 2020/10/25 07:57 by ebonnard

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