IGEM:IMPERIAL/2006/LabCalendar/2006-8-29

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Current revision (11:48, 30 August 2006) (view source)
(Ligations)
 
Line 29: Line 29:
**Digest DNA from Maxi prep stock
**Digest DNA from Maxi prep stock
**Run Maxi prep gel
**Run Maxi prep gel
 +
**[[Image:Maxi_j37024.pdf]]
**Glass Milk Purify insert and vector (J37023 and 1I respectively)
**Glass Milk Purify insert and vector (J37023 and 1I respectively)
**Leave to ligate overnight
**Leave to ligate overnight

Current revision



TO DO:

*Culture up the ligated parts (J37022 & J37025) for miniprep (to check whether ligations from Monday 
 were successful) from plates in incubator 
*Make frozen stock of 4G(S01656), J37018 and colony 2 of J37020
*Make more 50 times TAE
*Culture for testing: T9002, J37015, J37020 (colony 2)
*To order: LB medium (or make up ourselves & send to autoclave) & make some LB Amp to have in stock 
           Autoclaved flasks (and cuvettes?)
           Need more eppendorfs
           More LB Amp plates ? (How many more do we need for our remaining time - there are about 10 left)
*Gels from the weekend need to be analysed: Were J37018 & J37024 ligations successful ?

LoxP PCR

  • Run once more, this time with DMSO, and a larger amount of maxiprepped DNA
  • Major breakthrough - discovered that the reason for the repeated failures maybe that the DNA is there, but is not showing up.
  • Today's gel was looked at immediately after being run and two distinct bands could be seen - however these disappeared once left for a few minutes.
  • Once immersed in Ethidium bromide solution, the bands reappeared under the UV light.

Ligations

  • Repeat ligation J37023(Immunotagged AiiA)->1I
    • Digest DNA from Maxi prep stock
    • Run Maxi prep gel
    • Image:Maxi j37024.pdf
    • Glass Milk Purify insert and vector (J37023 and 1I respectively)
    • Leave to ligate overnight

Testing

Culture Up bacteria for testing tomorrow.

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