Haynes Lab:Notebook/CRISPR Editing/2015/01/27

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01/27/2015

Picked two colonies from KAH126 plate, put them into 2mLs of LB + amp. Let it shake overnight for mini preps tomorrow.



Luciferase assay on Luc14, GAL4-EED+dox, GAL4-EED 1-3

Wang lab has some old Taq polymerases that expired about 2 years ago. I'm testing them by amplifying g033 cloned into pX330 using the sequencing primers P139 and P140. If any of these polymerases work, I can use them to add dATPs to the ends of my Phusion PCR product for topo cloning.

Sample # Polymerase Buffer
12x taq mmN/A
22x taq mm*N/A
32x taq mm**N/A
4long amp taqLongAmp 5x buffer
5T4 DNA polymeraseStand taq Mg-free 10x buffer***
6Taq DNA polymeraseStand taq Mg-free 10x buffer

'*left this out for a long time, this one has more in it
'**left this out for a long time, this one has less in it
'***This is not the ideal buffer for this polymerase
Reactions 1-2

Thing amount
Taq MM 2x5
F primer0.3
R primer0.3
template0.3
H2O4

Reaction 4

Thing amount
LongAmp 5x buffer2
dNTPs0.3
F primer0.3
R primer0.3
template0.3
H2O6.5
long amp taq0.3

Reaction 5

Thing amount
Stand taq Mg-free 10x buffer1
dNTPs0.3
F primer0.3
R primer0.3
template0.3
H2O7.5
T4 DNA polymerase0.3

Reaction 6

Thing amount
Stand taq Mg-free 10x buffer1
dNTPs0.3
F primer0.3
R primer0.3
template0.3
H2O7.5
Taq DNA polymerase0.3



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