CDNA clean up - Revision history

128.111.227.84 at 15:56, 12 September 2012

2012-09-12T15:56:56Z

128.111.227.84 at 15:54, 12 September 2012

2012-09-12T15:54:59Z

128.111.227.84: Created page with "ds cDNA clean-up for Illumina sequencing (Agencourt AMPure XP) 1. Vortex beads to ensure paramagnetic beads are fully suspended 2. Transfer the samples to 1.5 mL tubes if necess…"

2012-09-12T15:54:09Z

Created page with "ds cDNA clean-up for Illumina sequencing (Agencourt AMPure XP) 1. Vortex beads to ensure paramagnetic beads are fully suspended 2. Transfer the samples to 1.5 mL tubes if necess…"

New page

ds cDNA clean-up for Illumina sequencing (Agencourt AMPure XP)

1. Vortex beads to ensure paramagnetic beads are fully suspended
2. Transfer the samples to 1.5 mL tubes if necessary
3. Add the appropriate volume of AMPure XP beads: multiply PCR volume by 1.8 (1.8 µL beads for 1.0 µL PCR product).
4. Mix by pipetting 10 times (color should be homogeneous). Let the mixed samples incubate at room temperature for 10 minutes. This step binds PCR products to the paramagnetic beads.
5. Transfer reactions to 0.2 mL tubes.
6. Place on the magnet plate for 2-5 minutes to separate beads from the solution. Check to make sure solution is clear before
7. Aspirate the clear solution without disturbing the beads
8. Dispense 200 µL of 70% EtOH to each tube and incubate at RT for 30 seconds. Aspirate out the ethanol and discard. Repeat for a total of two washes. Do not disturb the magnetic beads.
9. Incubate at RT for 5 minutes to dry the beads.
10. Off the magnetic plate, add 40 µL elution buffer and mix by pipetting 10 times. Elution is quite rapid and it is not necessary for the beads to go back into solution for it to occur.
11. Return tubes to the magnet plate and incubate at RT for 2-5 minutes.
12. Transfer the eluent to clean 1.5 mL tubes.

@@ Line 1: / Line 1: @@
 ds cDNA clean-up for Illumina sequencing (Agencourt AMPure XP)
-[1] Vortex beads to ensure paramagnetic beads are fully suspended
+# Vortex beads to ensure paramagnetic beads are fully suspended
-.	Transfer the samples to 1.5 mL tubes if necessary
+# Transfer the samples to 1.5 mL tubes if necessary
-.	Add the appropriate volume of AMPure XP beads: multiply PCR volume by 1.8 (1.8 µL beads for 1.0 µL PCR product).
+# Add the appropriate volume of AMPure XP beads: multiply PCR volume by 1.8 (1.8 µL beads for 1.0 µL PCR product).
-.	Mix by pipetting 10 times (color should be homogeneous). Let the mixed samples incubate at room temperature for 10 minutes. This step binds PCR products to the paramagnetic beads.
+# Mix by pipetting 10 times (color should be homogeneous). Let the mixed samples incubate at room temperature for 10 minutes. This step binds PCR products to the paramagnetic beads.
-.	Transfer reactions to 0.2 mL tubes.
+# Transfer reactions to 0.2 mL tubes.
-.	Place on the magnet plate for 2-5 minutes to separate beads from the solution. Check to make sure solution is clear before
+# Place on the magnet plate for 2-5 minutes to separate beads from the solution. Check to make sure solution is clear before
-.	Aspirate the clear solution without disturbing the beads
+# Aspirate the clear solution without disturbing the beads
-.	Dispense 200 µL of 70% EtOH to each tube and incubate at RT for 30 seconds. Aspirate out the ethanol and discard. Repeat for a total of two washes. Do not disturb the magnetic beads.
+# Dispense 200 µL of 70% EtOH to each tube and incubate at RT for 30 seconds. Aspirate out the ethanol and discard. Repeat for a total of two washes. Do not disturb the magnetic beads.
-.	Incubate at RT for 5 minutes to dry the beads.
+# Incubate at RT for 5 minutes to dry the beads.
-.	Off the magnetic plate, add 40 µL elution buffer and mix by pipetting 10 times. Elution is quite rapid and it is not necessary for the beads to go back into solution for it to occur.
+# Off the magnetic plate, add 40 µL elution buffer and mix by pipetting 10 times. Elution is quite rapid and it is not necessary for the beads to go back into solution for it to occur.
-.	Return tubes to the magnet plate and incubate at RT for 2-5 minutes.
+# Return tubes to the magnet plate and incubate at RT for 2-5 minutes.
-.	Transfer the eluent to clean 1.5 mL tubes.
+# Transfer the eluent to clean 1.5 mL tubes.