Omega Bio-Tek Mag-Bind SeqDTR M1300-05 User manual

Mag-Bind® SeqDTR™
M1300-05 5 mL
M1300-08 50 mL
M1300-50 500 mL
January 2013

1
Introduction and Overview.......................................................2
Illustrated Protocol........................................................................3
Kit Contents and Storage...........................................................4
Mag-Bind® SeqDTR™ 96-well Plate Protocol......................5
Mag-Bind® SeqDTR™ 384-well Plate Protocol.....................7
Troubleshooting Guide...............................................................9
Ordering Information................................................................10
Manual Revision: January 2013
Mag-Bind® SeqDTR™
Table of Contents

2
Introduction and Overview
Excess unincorporated, non-radioactive label can cause high background uorescence
in automated sequencing gels. For optimal sequencing results, remaining labeled
dideoxynucleotides should be removed prior to electrophoresis.
Omega Bio-tek’s Mag-Bind® SeqDTR™ is designed to eectively and reliably remove
unincorporated terminators from sequencing reactions. The system combines Omega
Bio-tek’s proprietary chemistries with the reversible nucleic acid-binding properties
of magnetic beads to eliminate excess nucleotides, primers, and small, non-targeted
amplication products such as primer dimers. This kit is designed for both manual and
fully automated purication of sequencing products.
The Mag-Bind® SeqDTR™ magnetic particles technology provides a better solution
for nucleic acid purication than centrifugation and vacuum-based technologies. The
product can be easily scaled up while providing simple user-friendly handling procedures.
If using the Mag-Bind® SeqDTR™ for the rst time, please read this booklet to become
familiar with the procedures. Sequencing products are mixed with the Mag-Bind®
SeqDTR™. DNA selectively binds to the Mag-Bind® SeqDTR™ particles. With one rapid
wash step, trace contaminants such as nucleotides, primers and small, non-targeted
amplication products are removed. Pure DNA is eluted in low salt buer or water.
Puried DNA can be directly used in downstream applications without the need for
further purication.
New in this edition:
• The manual has been updated to reect the product name change from the old
name of Mag-Bind® SE DTR to the new name of Mag-Bind® SeqDTR™.

3
Illustrated Protocol
Add Mag-Bind®SeqDTR™and 85% Ethanol, Mix
Magnetize and remove cleared supernatant
Wash 2 x with 85% Ethanol
Elute and transfer DNA to a new plate

4
Kit Contents and Storage
Storage and Stability
Mag-Bind® SeqDTR™ is stable for at least 9 months from the date of purchase when stored
at 2-8oC. Contents of the kit should never be frozen at any time.
Product No. M1300-05 M1300-08 M1300-50
Mag-Bind® SeqDTR™ 5 ml 50ml 500 ml
Preparations 500* / 1,000** 5,000* / 10,000** 50,000 */ 100,000**
User Manual P P P
* Based on a typical 10 μL reaction volume in a 96-well format
** Based on a typical 5 μL reaction volume in a 384-well format

5
Mag-Bind® SeqDTR™ - 96-well Plate Protocol
Mag-Bind®SeqDTR™96 Plate Format Protocol
Materials and Equipment to be Supplied by User:
• 85% ethanol (do not use denatured ethanol)
• Magnetic separation device compatible with 96-well PCR plates
• Multichannel pipet
• Reservoirs
• 96-well plate capable of being used in sequencers
• Elution Buer (Cat# PDR048 or 10 mM Tris pH 8.5, TE Buer, 0.1 mM EDTA, or diH₂O)
1. Thoroughly shake the Mag-Bind® SeqDTR™ to fully resuspend the magnetic beads.
2. Add 10 μL Mag-Bind® SeqDTR™ to each well.
Note: Use 10 μL Mag-Bind® SeqDTR™ regardless of the volume of the sequencing
reaction.
3. Add 85% ethanol according to table below and mix the sample thoroughly by
pipetting up and down 7-10 times.
Note: Do not use denatured ethanol. Always prepare fresh 85% ethanol within 3
days of use and store tightly capped.
Reaction volume (μL) 85% Ethanol (μL)
5 30
10 40
15 50
20 60
4. Place the plate on a magnetic separation device to magnetize the Mag-Bind®
SeqDTR™. Let sit at room temperature until the Mag-Bind® SeqDTR™is completely
cleared from solution.
5. Aspirate and discard the supernatant. Do not disturb the Mag-Bind® SeqDTR™.

6
6. Add 100 μL 85% ethanol to each well. It is not necessary to resuspend the Mag-Bind®
SeqDTR™.
7. Let sit at room temperature until the Mag-Bind® SeqDTR™is completely cleared from
solution.
8. Aspirate and discard the supernatant. Do not disturb the Mag-Bind® SeqDTR™.
9. Repeat Steps 6-8 for a second 85% ethanol wash step.
10. Leave the plate on the magnetic separation device for 10-15 minutes to air dry the
Mag-Bind® SeqDTR™. Remove any residue liquid with a pipettor.
Note: It is important to dry the Mag-Bind® SeqDTR™before elution. Residual ethanol
may interfere with downstream applications.
11. Add 40 μL Elution Buer (or 10 mM Tris pH 8.5, TE Buer, 0.1 mM EDTA, or diH2O) to
each well.
12. Pipet up and down 20 times to mix thoroughly.
13. Let sit at room temperature for 5 minutes.
14. Place the plate on a magnetic separation device to magnetize the Mag-Bind®
SeqDTR™. Let sit at room temperature until the Mag-Bind® SeqDTR™is completely
cleared from solution.
15. Transfer 30-35 µL cleared supernatant containing puried sequencing product to a
new plate capable of being used in sequencer.
Mag-Bind® SeqDTR™ - 96-well Plate Protocol

7
Mag-Bind®SeqDTR™- 384-well Plate Protocol
Additional Materials Supplied by User:
• 85% ethanol (do not use denatured ethanol)
• Magnetic separation device compatible with 384-well microplates
• Multichannel pipet
• Reservoirs
• 384-well plate capable of being used in sequencers
• Elution Buer (Cat# PDR048 or 10 mM Tris pH 8.5, TE Buer, 0.1 mM EDTA, or diH₂O)
1. Thoroughly shake the Mag-Bind® SeqDTR™ to fully resuspend the magnetic beads.
2. Add 5 μL Mag-Bind® SeqDTR™ to each well.
Note: Use 5 μL Mag-Bind® SeqDTR™ regardless of the volume of the sequencing
reaction.
3. Add 85% ethanol according to table below and mix the sample thoroughly by
pipetting up and down 7-10 times.
Note: Do not use denatured ethanol. Always prepare fresh 85% ethanol within 3
days of use and store tightly capped.
Reaction volume (μL) 85% Ethanol (μL)
5 14.3
10 21.4
15 28.6
4. Place the plate on a magnetic separation device to magnetize the Mag-Bind®
SeqDTR™. Let sit at room temperature until the Mag-Bind® SeqDTR™is completely
cleared from solution.
5. Aspirate and discard the supernatant. Do not disturb the Mag-Bind® SeqDTR™.
6. Add 30 μL 85% ethanol to each well. It is not necessary to resuspend the Mag-Bind®
SeqDTR™.
Mag-Bind® SeqDTR™ - 384-well Plate Protocol

8
7. Let sit at room temperature until the Mag-Bind® SeqDTR™is completely cleared from
solution.
8. Aspirate and discard the supernatant. Do not disturb the Mag-Bind® SeqDTR™.
9. Repeat Steps 6-8 for a second 85% ethanol wash step.
10. Leave the plate on the magnetic separation device for 10-15 minutes to air dry the
Mag-Bind® SeqDTR™. Remove any residue liquid with a pipettor.
Note: It is important to dry the Mag-Bind® SeqDTR™before elution. Residual ethanol
may interfere with downstream applications.
11. Add 15-20 μL Elution Buer (or 10 mM Tris pH 8.5, TE Buer, 0.1 mM EDTA, or diH2O)
to each well.
12. Pipet up and down 20 times to mix thoroughly.
13. Let sit at room temperature for 5 minutes.
14. Place the plate on a magnetic separation device to magnetize the Mag-Bind®
SeqDTR™. Let sit at room temperature until the Mag-Bind® SeqDTR™is completely
cleared from solution.
15. Transfer the cleared supernatant containing puried sequencing product to a new
plate capable of being used in sequencer.
Mag-Bind® SeqDTR™ - 384-well Plate Protocol

9
Troubleshooting Guide
Please use this guide to troubleshoot any problems that may arise. For further assistance,
please contact the technical support sta , toll free, at 800-832-8896.
Possible Problems and Suggestions
Problem Cause Solution
Dye terminator
remain in the
eluted DNA and
caused blobs
Supernatant is not
removed completely
Make sure to remove any liquid drops
from each well of the plate.
Too much BigDye® Use less BigDye® per reaction.
Insucient washing During steps 6-9, mix beads to wash
more eectively.
Problem Cause Solution
Low Signal
Ethanol concentration is
not correct
Make sure to use correct volume of
ethanol.
Low ethanol
concentration
Check the ethanol concentration, use
fresh ethanol if necessary.
Magnetic beads are lost
during the process
Make sure not to remove any magnetic
beads during aspiration.

10
Ordering Information
The following components are available for purchase separately.
(Call Toll Free at 1-800-832-8896)
Product Part Number
Nuclease-free Water (1 mL) PD092
Elution Buer (100 mL) PDR048
Multichannel Disposable Reservoirs (100/pk) AC1331-01
96-well Microplate (500 µL) (25/pk) EZ9604-02
Mag-Bind® SeqDTR™ (50 mL) M1300-08
Mag-Bind® SeqDTR™ (500 mL) M1300-50
Mag-Bind® E-Z Pure (50 mL) M1380-01
Mag-Bind® E-Z Pure (500 mL) M1380-02
Mag-Bind® and SeqDTR™ are trademarks of Omega Bio-tek, Inc.
BigDye is a registered trademarks of Applied Biosystems or its subsidiaries in the U.S
PCR is a patented process of Homan-La Roche. Use of the PCR process requires a license.

11
Notes:

12
Notes: