AlanFung:LabNotes/Capturing/2014-1-23: Difference between revisions

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Line 18: Line 18:
| height="13" | ppMALAT1
| height="13" | ppMALAT1
| align="center" | 3036
| align="center" | 3036
|style="background-color:#EBF1DE" align="center" | 0.99
|style="background-color:#EBF1DE" align="center" | 0.40
| align="center" | 100
| align="center" | 20
| align="center" | 30
| align="center" | 60
|style="background-color:#EBF1DE" align="center" | 29.01
|style="background-color:#EBF1DE" align="center" | 59.60


|-  align="center"
|-  align="center"
| height="13" | MALAT1 Template
| height="13" | MALAT1 Template
| align="center" | 1485
| align="center" | 1485
|style="background-color:#EBF1DE" align="center" | 1.35
|style="background-color:#EBF1DE" align="center" | 0.40
| align="center" | 100
| align="center" | 20
| align="center" | 20
|style="background-color:#EBF1DE" align="center" | 18.65
| align="center" | 30
|style="background-color:#EBF1DE" align="center" | 29.6


|-  align="center"
|-  align="center"
| height="13" | ppillumina_155
| height="13" | ppillumina_155
| align="center" | 3267
| align="center" | 3267
|style="background-color:#EBF1DE" align="center" | 0.92
|style="background-color:#EBF1DE" align="center" | 0.40
| align="center" | 100
| align="center" | 20
| align="center" | 30
| align="center" | 65
|style="background-color:#EBF1DE" align="center" | 29.08
|style="background-color:#EBF1DE" align="center" | 64.6


|-  align="center"
|-  align="center"
| height="13" | 10BP
| height="13" | 10BP
| align="center" | 1000
| align="center" | 1000
|style="background-color:#EBF1DE" align="center" | 0.10
|style="background-color:#EBF1DE" align="center" | 0.40
| align="center" | 100
| align="center" | 20
| align="center" | 1
| align="center" | 20
|style="background-color:#EBF1DE" align="center" | 99
|style="background-color:#EBF1DE" align="center" | 19.6


|}
|}

Revision as of 20:26, 23 January 2014

Capturing ppMALAT1 template w ppMALT1 ordered form IDT

TBU GEL to verify probe template size

  • Aim for 100ng/band
  • 10bp Ladder
  • ppMALAT1 (92nt, 100uM=3036ng/ul)
  • MALAT1 Template (45nt, 100uM=1485ng/ul)
  • ppillumina_155 (99nt, 100uM=3267ng/ul)
Sample ng/ul ul ng/ul ul H2O
ppMALAT1 3036 0.40 20 60 59.60
MALAT1 Template 1485 0.40 20 30 29.6
ppillumina_155 3267 0.40 20 65 64.6
10BP 1000 0.40 20 20 19.6
  • Pre-run the gel for 30 min at 200V
  • Dilute sample to 20ng/uL
  • Denature samples and ladder at 75C for 7 min, then quickly transfer to cooling rack, wait for 5 mins
  • Flush the wells very well with P1000 & P200 pipette to remove all urea.
  • NOTE: DO NOT TOUCH THE TUBE WITH YOUR HANDS IN THE FOLLOWING STEP, GRAB THE CAP
  • Load the samples into the wells, run at 200V for 30 min.
  • Stain gel with 3uL SYBR Gold in ~100 mL of TBE in clean tray.
  • Place tray on an orbital shaker for ~5 min.

ssDNA Qubit to quantify probe and template

Probe calculations ppMALAT1