GALENVS SPR0016 Plant RNA Extraction Kit User Guide

GALENVS SPR0016 Plant RNA Extraction Kit

Product Information

The Plant RNA Extraction Kit is designed for efficient extraction of RNA from plant samples. It includes all the necessary reagents and components for the extraction process.

Product Usage Instructions

Part RNA Extraction

1. Add up to 50mg of fresh plant leaves sample to the lysis bead tube provided.
2. Mix for 10 mins using TissueLyser at max speed or vortex for 10 mins; then centrifuge at 20,000g for 2 mins.
3. Add Binding Buffer #1 (columns 1 & 7). You can add up to 16 samples.
4. Place the plate into the miQron, taking care that the label is facing outward.
5. Insert two combs.
6. Select PRKit – Part A and press “Run”.
7. When the program is complete, remove the plate from miQron and discard the combs.

Part DNase Treatment

1. Mix DNase with Reaction Buffer in a microfuge tube by gently inverting the tube a few times.
2. Add DNase to Elution Buffer (columns 6 & 12) in the RNA Extraction Kit Plate. Dispense directly into the elution buffer, not on the walls.
3. Gently shake the plate for 10 seconds by hand and incubate at room temperature for 20 mins.
4. Transfer 100uL of the Elution Buffer (columns 6 & 12) from the RNA Extraction Kit Plate to each well (columns 1 & 7) of the RNA Extraction Kit Plate.
5. Place the plate into the miQron, taking care that the label is facing outward.
6. Insert two combs.
7. Select PRKit – Part B and press “Run”.
8. When the program is complete, remove the plate from miQron and discard the combs.

Columns 5 and 11 of the RNA Extraction Kit Plate contain the purified RNA elution.

RNA Extraction

• Binding Buffer (600µl)
  • Columns 1 & 7
• Wash #1 Buffer (600µl)
  • Columns 2 & 8, 3 & 9
• Wash #2 Buffer (600µl)
  • Columns 4 & 10, 5 & 11
• Elution Buffer (100µl)
  • Columns 6 & 12
• 400µl sample added to Binding Buffer
• 100µl extracted and purified RNA elution
• 400µl sample added to Binding Buffer
• 100µl extracted and purified RNA elution

PRKit–A miQron protocol parameters

Step Name| Column| Volume (µl)| Time (sec)| Mixing Speed (1–10)| Dry Time (sec)| Magnet Capture Time (sec)
---|---|---|---|---|---|---
Binding| 1 & 7| 600| 300| 7| 0| 150
Wash #1| 2 & 8| 600| 60| 7| 0| 90
Wash #1| 3 & 9| 600| 60| 7| 0| 90
Wash #2| 4 & 10| 600| 60| 7| 0| 90
Wash #2| 5 & 11| 600| 60| 7| 300| 90
Elution| 6 & 12| 100| 60| 10| 0| 150
Discard Comb| 2 & 8| 600| 0| 5| 0| 0

Using Instructions

1. Add up to 50mg of fresh plant leaves sample to the lysis bead tube provided.
2. Add 600µl Lysis Buffer, and 60µl PPB. Mix for 10 mins using TissueLyser at max speed or vortex for 10 mins; then centrifuge at 20,000g for 2 mins.
3. To the RNA Extraction Kit Plate A transfer up to 400µl of supernatant to Binding Buffer #1 (columns 1 & 7). You can add up to 16 samples.
4. Place plate into the miQron, taking care that the label is facing outward.
5. Insert two combs.
6. Select PRKit – Part A and press
   • When program is complete, remove plate from miQron and discard combs.

DNase Treatment

• Binding Buffer (400µl)
  • Columns 1 & 7
• Functionalized Beads (200µl)
  • Columns 2 & 8
• Wash #3 Buffer (600µl)
  • Columns 3 & 9
• Elution Buffer (50µl)
  • Columns 5 & 11
• 100µl sample added to Binding Buffer
• 50µl extracted and purified RNA elution
• 100µl sample added to Binding Buffer
• 50µl extracted and purified RNA elution

PRKit-B miQron protocol parameters

Step Name| Column| Volume (µl)| Time (sec)| Mixing Speed (1–10)| Dry Time (sec)| Magnet Capture Time (sec)
---|---|---|---|---|---|---
Bead Transfer| 2 & 8| 200| n/a| n/a| 0| 100
Binding| 1 & 7| 400| 12| 7| 0| 110
Wash #3| 3 & 9| 600| 12| 6| 0| 100
Elution| 5 & 11| 50| 12| 7| 0| 200
Discard Comb| 3 & 9| 600| 0| 5| 0| 0

Using Instructions

• Add 100µl of DNase Reaction Buffer to DNase pellet and then add 100µl of glycerol.
  • Mix by gently inverting the tube.
  • Reconstituted pellet must be stored at -20 °C.
• For each RNA sample, prepare DNase Buffer by adding 10µl of DNase prepared in the previous step to 40µl of DNase Reaction Buffer in a microfuge tube.
  • Mix DNase with buffer by gently inverting the tube a few times.
• To the RNA Extraction Kit Plate A add 50µl of DNase Buffer to Elution Buffer (columns 6 & 12). Dispense directly into the elution buffer not on the walls. If there are any droplets of elution buffer left on the walls, slide them to the well bottom by gently shaking or tapping the plate on a solid surface.
• Gently shake Plate A for 10 seconds by hand and incubate at room temp for 20 mins. From the RNA Extraction Kit Plate A transfer 100uL of the Elution Buffer (columns 6 & 12) to each well (columns 1 & 7) of the RNA Extraction Kit Plate B.
• Place Plate B into the miQron, taking care that the label is facing outward.
• Insert two combs.
• Select PRKit – Part B and press
  • When program is complete, remove plate from miQron and discard combs.
  • Columns 5 and 11 contain the purified RNA elution.

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