Haynes Lab:Notebook/CRISPR Editing/2015/01/07: Difference between revisions

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(Autocreate 2015/01/07 Entry for Haynes_Lab:Notebook/CRISPR_Editing)
 
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==07//2014==
==01/07/2015==
 
Set up 2-50μL PCR reactions for untreated Luc14 gDNA and a subset of treated Luc14.<br>
<br>


{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Mastermix 1'''
| align="center" style="background:#f0f0f0;"|''''''
| align="center" style="background:#f0f0f0;"|''''''
|-
| ||1rxn||28.3rxns
|-
| H20||30||849
|-
| HF 5X MasterMix||10||283
|-
| dNTPs||1||28.3
|-
| FP||2.5||70.75
|-
| RP||2.5||70.75
|-
| ||46||1301.8
|-
| Mastermix 1||||
|-
| ||1rxn||2.1rxns
|-
| MM1||46||96.6
|-
| Template||2||4.2
|-
| DMSO||1.5||3.15
|-
| Phusion||0.5||1.05
|-
| ||50||205
|}
Transfer 50μL of MM2 into each of 4-PCR tubes.<br>
PCR
:98°C for 3 min
:36 cycles
::98°C for 10s
::66°C for 30s
::72°C for 60s
:72°C for 10 min
:4°C forever
<br>
'''Sample g039 may have too much gDNA, samples g045 and g046 were in non-autoclaved tubes, sample g046 did not have 50ul for each tube'''


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01/07/2015

Set up 2-50μL PCR reactions for untreated Luc14 gDNA and a subset of treated Luc14.

Mastermix 1 ' '
1rxn 28.3rxns
H20 30 849
HF 5X MasterMix 10 283
dNTPs 1 28.3
FP 2.5 70.75
RP 2.5 70.75
46 1301.8
Mastermix 1
1rxn 2.1rxns
MM1 46 96.6
Template 2 4.2
DMSO 1.5 3.15
Phusion 0.5 1.05
50 205

Transfer 50μL of MM2 into each of 4-PCR tubes.
PCR

98°C for 3 min
36 cycles
98°C for 10s
66°C for 30s
72°C for 60s
72°C for 10 min
4°C forever


Sample g039 may have too much gDNA, samples g045 and g046 were in non-autoclaved tubes, sample g046 did not have 50ul for each tube


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