Difference between revisions of "SDS Page-Gel for all the bovine casein inserts"
Emi Nikolov (talk | contribs) |
Emi Nikolov (talk | contribs) |
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'''Notes:''' Emi | '''Notes:''' Emi | ||
'''Aims:''' | '''Aims:''' Confirmation of bovine casein inserts from liquid culture of yeast transformed cells | ||
'''Materials''' | '''Materials''' | ||
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Cytosin | Cytosin | ||
Transformed negative control | |||
Bov alpha s1 (1) Transformed liqid culture | |||
Bov beta B (8) Transformed liquid culture | |||
Bov alpha S2 (3) Transformed liquid culture | |||
Bov beta A2 (6B) Transformed liquid culture | |||
Bov kappa (7F) Transformed liquid culture | |||
Chymosin + casein (Chymosin cuts casein and separates it to 2 bands) | |||
Ladder | |||
| Line 30: | Line 48: | ||
Centrifuge the falcon tubes, 5 minutes 2800xg | Centrifuge the falcon tubes, 5 minutes 2800xg | ||
Take 500ul of distilled water in eppendorf tube + 1 tiny drop of casein | Take 500ul of distilled water in eppendorf tube + 1 tiny drop of casein (use 200ul hp take it) - 2times | ||
Take 15ul of chymosin +1 tiny drop of casein in 250ul of water | Take 15ul of chymosin +1 tiny drop of casein in 250ul of water | ||
Take 250ul of 7F (Bov Kappa) + 15ul of | Take 250ul of 7F (Bov Kappa) + 15ul of Chymosinin | ||
Take 250ul of water and 15ul of chymosin | |||
Take 40ul of 1, 8, 3B, 6B, +7F also the negative control to eppendorf tubes | Take 40ul of 1, 8, 3B, 6B, +7F also the negative control to eppendorf tubes | ||
| Line 40: | Line 59: | ||
Preparation fo 20x running buffer | Preparation fo 20x running buffer | ||
Pour | Pour 50ul of 10x running buffer; make it to 450ul by adding the distilled water - total 500ul | ||
20ul of loading dye to each tubes except casein; chymosin and casein; chymosin and 7F | |||
Thus we have to add total 120ul of dye | |||
Boil all the tubes for 10 mins at 95C (inorder to denature the protein structure to linear) | |||
(only | |||
Revision as of 22:28, 17 January 2015
17 January 2015: SDS Page-Gel All 5 Bovine Casein Inserts
Location: BioCurious
Participants: Lafia, Johan, Meenakshi, Emi, Rachel
Notes: Emi
Aims: Confirmation of bovine casein inserts from liquid culture of yeast transformed cells
Materials
Gel loading tips
Tris-Acelate SDS Running Buffer novex life tech
Bio-Rad pipette tips
Native gels... (need details)
ladder 20x
Casein
Cytosin
Transformed negative control
Bov alpha s1 (1) Transformed liqid culture
Bov beta B (8) Transformed liquid culture
Bov alpha S2 (3) Transformed liquid culture
Bov beta A2 (6B) Transformed liquid culture
Bov kappa (7F) Transformed liquid culture
Chymosin + casein (Chymosin cuts casein and separates it to 2 bands)
Ladder
Protocol
Centrifuge the falcon tubes, 5 minutes 2800xg
Take 500ul of distilled water in eppendorf tube + 1 tiny drop of casein (use 200ul hp take it) - 2times
Take 15ul of chymosin +1 tiny drop of casein in 250ul of water
Take 250ul of 7F (Bov Kappa) + 15ul of Chymosinin Take 250ul of water and 15ul of chymosin
Take 40ul of 1, 8, 3B, 6B, +7F also the negative control to eppendorf tubes
Preparation fo 20x running buffer
Pour 50ul of 10x running buffer; make it to 450ul by adding the distilled water - total 500ul
20ul of loading dye to each tubes except casein; chymosin and casein; chymosin and 7F
Thus we have to add total 120ul of dye
Boil all the tubes for 10 mins at 95C (inorder to denature the protein structure to linear) (only