Matt:LabNotes/2014-5-27: Difference between revisions
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>Mzcai (Created page with "==Artifical Rolonies== ===Procedure=== #Add 140ul 100nM MALAT1_template to each well #*Pre-heat to 85C #*Incubate @RT for 30 min #**C4 & C5 Incubate @RT for '''15 min''' #Aspi...") |
>Mzcai mNo edit summary |
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#Add 140ul 100nM MALAT1_template to each well | #Add 140ul 100nM MALAT1_template to each well | ||
#*Pre-heat to 85C | #*Pre-heat to 85C | ||
#*Incubate @RT for | #*Incubate @RT for '''75 min''' | ||
#**C4 & C5 Incubate @RT for ''' | #**C4 & C5 Incubate @RT for '''60 min''' | ||
#Aspirate | #Aspirate | ||
#Capture with | #Capture with 100pM ppMALAT1 | ||
#*Incubate @45C for 24 hours ( | #*Incubate @45C for 24 hours (6:30p- ) | ||
{| {{table}} | {| {{table}} | ||
| align="center" style="background:#f0f0f0;"|'''Component''' | | align="center" style="background:#f0f0f0;"|'''Component''' | ||
Line 14: | Line 14: | ||
| 10X Ampligase Buffer||80 | | 10X Ampligase Buffer||80 | ||
|- | |- | ||
| | | 2nM ppMALAT1||40 | ||
|- | |- | ||
| 5 U/ul Ampligase||80 | | 5 U/ul Ampligase||80 | ||
|- | |- | ||
| H2O|| | | H2O||600 | ||
|- | |- | ||
| Total||800 | | Total||800 | ||
Line 68: | Line 68: | ||
===Procedure=== | ===Procedure=== | ||
*Strip with 80% formamide for 15min @RT | *Use pre-made artificial MALAT1 rolonies from [[Matt:LabNotes/2014-5-16]] | ||
*Wash with ddH2O twice | **Strip with 80% formamide for 15min @RT | ||
**Wash with ddH2O twice | |||
'''Experimental & Negative Control''' (Positions 3-12) | '''Experimental & Negative Control''' (Positions 3-12) |
Revision as of 01:40, 28 May 2014
Artifical Rolonies
Procedure
- Add 140ul 100nM MALAT1_template to each well
- Pre-heat to 85C
- Incubate @RT for 75 min
- C4 & C5 Incubate @RT for 60 min
- Aspirate
- Capture with 100pM ppMALAT1
- Incubate @45C for 24 hours (6:30p- )
Component | Volume |
10X Ampligase Buffer | 80 |
2nM ppMALAT1 | 40 |
5 U/ul Ampligase | 80 |
H2O | 600 |
Total | 800 |
Exo I/III Test
Procedure
- Use pre-made artificial MALAT1 rolonies from Matt:LabNotes/2014-5-16
- Strip with 80% formamide for 15min @RT
- Wash with ddH2O twice
Experimental & Negative Control (Positions 3-12)
Component | Volume |
10X Ampligase Buffer | 50 |
2uM ppMALAT1 | 2.5 |
H2O | 447.5 |
Total | 500 |
Positive Control (Positions 1-2)
Component | Volume |
10X Ampligase Buffer | 10 |
H2O | 90 |
Total | 100 |