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Bleaching and other notes

Bleaching

Making 10ml bleach:

  • 3ml Sodium hypochloride
  • 1.5ml 5M KOH
  • 5.5ml dH20 or ddH20

Two types:

  1. Spot bleaching
  2. Traditional bleaching

For traditional bleaching:

bleaching.pdf

Need lots of worms - otherwise you risk losing them in the process…

Finish by removing excess liquid after overnight till just under 0.1ml line –> pipette the remaining liquid up and down 3 times before transferring the liquid onto the the non-contaminated 6cm plate

For spot bleaching:

  1. Add 50-100ul bleach to the corner of the uncontaminated plate away from the bacterial lawn
  2. Pick up 5-6 gravid worms and transfer onto bleach
  3. Shake plate a bit to separate worms
  4. Incubate O/N at 20C

Notes:

  • Can keep bleach for up to 1 week at 4C - use fresh wherever possible and especially if correct synchronization is desired
  • Synchronization:
    • Synchronization means worms hatch as larva starved and begin at the same point of growth after being fed
    • If bleach not 100% successful then this desynchronizes the worms
    • If synchronization is not needed then it's ok if not 100% - just to get rid of fungus and bacteria
  • Amount of bleaching done at one time recommended = 6 different strains or less - otherwise difficult to keep track of all of them
  • Filter tips are more useful to minimize cross-contamination

Other notes

  • Healthy chunks grown over the weekend will need rechunking on Monday - fast growth!
  • Try not to pick old worms as these have no more embryos
  • Pregnant worms looks like it has spots in the belly - hermaphrodite can house 300 eggs at a time

  • Transgenic - two types
    • EX array - can be lost kind of like a plasmid (no selection for it) - different markers to identify it:
      • Fluorescence
      • Multi-vulva defect –> EX carriers can rescue this defect
    • IS (Integrated Strain) - all worms have it
  • Pros and cons of EX or IS depend on the type of experiment you're doing
wiki/documentation/wiki/documentation/nicolina/bleaching_and_other_notes.txt · Last modified: 2019/11/14 17:39 by nzjacic

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