Difference between revisions of "UW-Stout/Nitrogen starvation FA19"

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(Created page with "Materials: Materials Nitrogen + Nitrogen - Histidine (x100) 1 ml 1ml Leucine (x100) 1ml 1ml Uracil (x100) 1ml 1ml CSM 65 mg 65mg YBN 171mg 171mg Glucose (40% w/v) 5ml 5ml Am...")
 
 
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Materials:
+
[[File:Yeast_Tourture_Ex_C+T1.jpg|1,700 × 2,200 |Caption]]
 
+
[[File:Yeast_Tourture_Ex._C+T2.jpg|1,700 x 2,200 |Caption]]
Materials
+
[[File:Nitrogen_Starvation_BY4735.jpg|463 x 599|Caption]]
Nitrogen +
 
Nitrogen -
 
Histidine (x100)
 
1 ml
 
1ml
 
Leucine (x100)
 
1ml
 
1ml
 
Uracil (x100)
 
1ml
 
1ml
 
CSM
 
65 mg
 
65mg
 
YBN
 
171mg
 
171mg
 
Glucose (40% w/v)
 
5ml
 
5ml
 
Ammonium Sulfate
 
0
 
500mg
 
Water
 
92ml
 
92ml
 
 
 
Equipment:  
 
Molecular Devices SpectraMax Plus 384 Microplate Reader
 
 
 
Procedure:
 
 
 
Part 1
 
Label two seperate 250ml glass media jars with Nitrogen + and Nitrogen -
 
In Nitrogen + media jar mix in correct portions from above materials list
 
Histidine (x100)
 
Leucine (x100)
 
Uracil (x100)
 
CSM
 
YBN
 
Water
 
In Nitrogen - Media jar mix in correct portions form above materials list
 
Histidine (x100)
 
Leucine (x100)
 
Uracil (x100)
 
CSM
 
YBN
 
Ammonium Sulfate
 
Water
 
Mix with stirring for 10-15 minutes
 
Place both glass media jars in plastic bin with autoclave tape secured around a loosened lid, then proceed to autoclave at 121°C for 15 minute
 
With autoclave gloves on retrieve the solutions
 
When both media jars are cool, mix in 5 ml of glucose to both Nitrogen + and Nitrogen - solutions
 
 
 
Part 2
 
Gather all yeast culture media
 
In Biosafety cabinet pipet 1ml of each culture twice into separate microcentrifuge tubes
 
Centrifuge tubes for 30 sec at 5000 rcf
 
In biosafety cabinet aspirate supernatant into beaker of bleach.
 
Label tubes with 1 nitrogen - and 1 nitrogen + for each yeast culture
 
Add 95ml of solution corresponding to the label
 
Vortex solutions
 
Pipet all of the solution into wells on the microplate
 

Latest revision as of 23:23, 17 December 2019

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