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Showing posts with label media. Show all posts
Showing posts with label media. Show all posts
Wednesday, January 25, 2023
Monday, October 26, 2015
use nutrient plates for yeast
Oct 26-Nov3. 2015
Use nutrient plates for yeast. The growth is extremely slow. Small colonies can be seen after 1 week.
Use nutrient plates for yeast. The growth is extremely slow. Small colonies can be seen after 1 week.
Wednesday, October 29, 2014
Monday, September 8, 2014
YPD plates
For 1000 ml
YE 10 g (Difco)
Peptone 10g (Bacto)
Agar 20 g
fill to 900 ml
weight at 1519gram
autoclave, 45min (autocalve a 100ml cylinder )
weigt at 1648gram.
add 50gram water.
20% glucose 100ml (for 2%glucose) (use the autoclaved cylinder)
http://openwetware.org/wiki/YPD
Monday, February 10, 2014
BIO125, FOA plate media and western blot media
=> For FOA plates, the media is -HIS-TRP-LEU-THR+URA
-LEU -THR for URA3's LEU2 promoter to be active. The presence of Leucine and Theorine lead to 10 fold drop of LEU2 expression.
-HIS to keep pMSH2
-TRP to keep pSH44 reporter plasmid
+URA for FOA assay, because we are selecting for mutant URA3.
So the FOA media is HIS-TRP-LEU-THR + URA
However, before the FOA plate, yeast cells should be grown in - URA to ensure intact dinucleotide repeat in pSH44.
So the regular media is -URA-HIS-TRP (-LEU-THR)
See Gammie04CBE
=> For MSH2 expression:
Gammie07,
Three type of pMSH2 was mentioned. pMSH2, pGBD-MSH2, and pGAL-MSH2.
The pMSH2 is under endogenous promoter. So, the media should be -His to keep pMSH2.
The pGBD-MSH2 was designed to have a constitutive expression of MSH2 protein with HA tag. This plasmid seems to be used for two-hybrid.
The pGAL-MSH2 contains a GAL10 promoter.
References
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC294742/pdf/jbacter00572-0372.pdf
-LEU -THR for URA3's LEU2 promoter to be active. The presence of Leucine and Theorine lead to 10 fold drop of LEU2 expression.
-HIS to keep pMSH2
-TRP to keep pSH44 reporter plasmid
+URA for FOA assay, because we are selecting for mutant URA3.
So the FOA media is HIS-TRP-LEU-THR + URA
However, before the FOA plate, yeast cells should be grown in - URA to ensure intact dinucleotide repeat in pSH44.
So the regular media is -URA-HIS-TRP (-LEU-THR)
See Gammie04CBE
=> For MSH2 expression:
Gammie07,
Three type of pMSH2 was mentioned. pMSH2, pGBD-MSH2, and pGAL-MSH2.
The pMSH2 is under endogenous promoter. So, the media should be -His to keep pMSH2.
The pGBD-MSH2 was designed to have a constitutive expression of MSH2 protein with HA tag. This plasmid seems to be used for two-hybrid.
The pGAL-MSH2 contains a GAL10 promoter.
References
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC294742/pdf/jbacter00572-0372.pdf
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