View
0
Download
0
Category
Preview:
Citation preview
MagnaMedics
Move to SimplicityHowTo: Automate MagSi-NGSPREP
on a PrimaDiag workstation
MagSi-NGSPREP automation protocol for PrimaDiag
This HowTo describes the preparations needed for running automated protocols with MagSi-NGSPREP on PrimaDiag liquid handling platforms.
Magnetic bead based MagSi-NGSPREP offers an efficient solution for clean-up and size selection in NGS library preparation. The simple and flexible protocols can be adjusted to your specific application and NGS platform.
All 4 protocols described in the MagSi-NGSPREP product manual have been automated for the PrimaMPS 46and PrimaRWS 46 liquid handling platform and are available for download. Protocols for other system configurations are available on request - please contact our technical support.isFor automation MagSi-NGSPREP one of the following platforms are required and available at yourPrimaDiag distributor:
PrimaMPS 46
Product Discription Quantity Catalog number
RWS 46 base unit Platform 40 x 60 cm – allows 1 or 2 Z-axis 1 10220
15 SBS deck 46 MPS 15 standard positions deck for PrimaMPS 46 1 10020
Z-axis Z-axis able to embed a pipetting head 1 10300
P8V250 8-channel pipeting head – 1-250µl (position Z1) 1 20285
PCII full1x PrimaControllerII license with the control computer 1 30100
TipsHolder250 Holder for tipbox 250µl 7 60420
Magnetic Working Unit Active magnetic unit for beads technology 1 40050
PrimaRWS 46
Product Discription Quantity Catalog number
RWS 46 base unit Platform 40 x 60 cm – allows 1 or 2 Z-axis 1 10220
16 SBS deck 46 16 standard positions deck for PrimaRWS 46 1 10020
Z-axis Z-axis able to embed a pipetting head 1 10300
P8V250 8-channel pipeting head – 1-250µl (position Z1) 1 20285
PCII full1x PrimaControllerII license with the control computer 1 30100
TipsHolder250 Holder for tipbox 250µl 7 60420
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 1
MagnaMedicsMove to Simplicity
Procedure:
Step 1. Import your protocolThe latest protocol files for your application are available on magnamedics.com/download/softwareAll protocol files are compatible with PrimaController II v.1.7.8.2 and above.
Protocol number in MagSi-NGSPREP manual
Protocol MagSi-NGSPREP Filename
4.1 Clean-up of enzymatic reaction MagSi-NGSprepCleanup_vX.X.zip
4.2 Left Side Size Selection MagSi-NGSprepLeftSideSizeSelection_vXX.zip
4.3 Right Side Size Selection MagSi-NGSprepRightSideSizeSelection_vX.X.zip
4.4 Double Sided Size Selection MagSi-NGSprepDoubleSidedSizeSelection_vX.X.zip
X.X notes the latest version number
1. Open the downloaded file. When opening the file you might get prompted for a password. If the password is not known, please contact MagnaMedics or your distributor. Copy the protocol file for the desired application to the computer running the PrimaController II software.
2. Open the PrimaController II software.
3. Click 'File' in the upper lefthand corner and choose 'Open project', followed by 'Browse'
4. Navigate to the application protocol file (.prws) and click 'Open'.
To offer the user the flexibility required for different applications a number of variables have been implemented, including sample volume, sample positions and desired MagSi-NGSPREP bead ratio. Prior to running the protocols these variables are shown and can be set to user needs.
5. Click the check mark at the bottom of the screen
6. Edit the variables to your preferences.
7. When all variables have been set, click the refresh button.
The amount of reagents needed for the protocol is recalculated and shown in the bottom part of the screen. Continue with the preperation of the deck.
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 2
MagnaMedicsMove to Simplicity
ProtocolMagSi-NGSPREP
Variable Min Max Description
4.1 Clean-upof enzymatic
reactions
Sample Volume (μl) 10 70 The volume of the samples
First Column withSamples
1 12 First column of samples to be processed
Last Column withSamples 1 12 Last column of samples to be processed
Ratio MagSi-NGSPREP:sample 1.0 1.8
Should be 1.8 for PCR cleanup. Alternatively,use the volume of beads recommended by
your library prep kit manual.
Elution Volume (μl) 35 - Volume of the final eluate
4.2 Left SideSize Selection
Sample Volume (μl) 50 70 The volume of the samples
First Column withSamples 1 12 First column of samples to be processed
Last Column withSamples
1 12 Last column of samples to be processed
Left Side Ratio 0.4 1.5The beads ratio corresponding with the
desired left side cutoff size.
Elution Volume (μl) 35 - Volume of the final eluate
4.3 Right SideSize Selection
Sample Volume (μl) 50 70 The volume of the samples
First Column withSamples
1 6 First column of samples to be processed
Last Column withSamples 1 12 Last column of samples to be processed
Right Side Ratio 0.4 1.5 The beads ratio corresponding with thedesired right side cutoff size.
Elution Volume (μl) 35 - Volume of the final eluate
4.4 DoubleSided SizeSelection
Sample Volume (μl) 50 70 The volume of the samples
First Column withSamples 1 12 First column of samples to be processed
Last Column withSamples
1 12 Last column of samples to be processed
Right Side Ratio 0.4 1.5The beads ratio corresponding with the
desired right side cutoff size.
Left Side Ratio 0.4 1.5 The beads ratio corresponding with thedesired left side cutoff size.
Elution Volume (μl) 35 - Volume of the final eluate
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 3
MagnaMedicsMove to Simplicity
Step 2. Prepare the deck of your platform
The protocols obtained from MagnaMedics are defined with the following deck layout. If you want to use adifferent deck layout please contact the MagnaMedics support team to request a protocol custom made for your needs.
1. Pipet tips2. Pipet tips3. Pipet tips4. Pipet tips5. Pipet tips *6. Liquid waste position7. Reagent position8. Pipet tips *9. Pipet tips *10. Sample output position11. Sample position (for PrimaRWS 46)12. Empty13. Empty14. Empty
MWU. Sample position
Positions marked with an asterisk are only applicable for the Left Side Size selection and clean-up protocols.
A) Labware
The following consumables are used in the predefined protocols. If you want to use different consumablesplease contact the MagnaMedics support team to request a protocol custom made for your needs.
Position Consumable Cat. #
Pipet tips 1-250 µl
Pipet tips Primadiag Tips250Primadiag Tips250Refill
8030080302
Microtiterplate
Sample position Axygen 96 PCR well plateMagnaMedics MM-Separator 96 SBS*
PCR-96-FS-CMD90005
Reagent position Primadiag Reservoir12 60301
Sample output position Thermo 96-Well Semi-Skirted AB-1100
Liquid waste position Primadiag DPW2mlsquare 60302
Waste PrimaDiag Waste slider
* Only applicable when using the PrimaRWS 46 platform
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 4
MagnaMedicsMove to Simplicity
B) Reagent position
The reagents need to be present in the vertical slots according to the table below. Please note that thelayout can be slightly different for a different protocol. It is not necessary to calculate the requiredvolumes by hand, the software presents the outcome of these calculations prior to running the protocol.
ProtocolMagSi-NGSPREP
Containercontent
Containerslot
Total required volume
4.1 Clean-up
MagSi-NGSPREP 1 (Sample volume (µl) * 1.8) * Number of samples + 200 µl
Ethanol 70% 2 180 * Number of samples + 200 µl
Ethanol 70% 3 180 * Number of samples + 200 µl
Elution buffer 4 Elution Volume (µl) * Number of samples + 200 µl
4.2 Left SideSize Selection
MagSi-NGSPREP 1 (Sample volume (µl) * Ratio) * Number of samples + 200 µl
Ethanol 85% 2 180 * Number of samples + 200 µl
Ethanol 85% 3 180 * Number of samples + 200 µl
Elution buffer 4 Elution Volume (µl) * Number of samples + 200 µl
4.3 Right SideSize Selection
MagSi-NGSPREP 1 (Sample volume (µl) * 1.8) * Number of samples + 200 µl
Ethanol 85% 2 180 * Number of samples + 200 µl
Elution buffer 4 Elution Volume (µl) * Number of samples + 200 µl
4.4 DoubleSided SizeSelection
MagSi-NGSPREP 1(Sample volume (µl) * Left Ratio) * Number of samples +
200 µl
Ethanol 85% 2 180 * Number of samples + 200 µl
Elution buffer 4 Elution Volume (µl)* Number of samples + 200 µl
Note: the 200 µl aliquot mentioned in all formula's is to account for dead volumes
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 5
MagnaMedicsMove to Simplicity
C) Sample position:
The sample plate to be processed is placed on the Magnetic Working Unit. To prevent the loss of reagents, the samples in the plate should be positioned in colums of 8.
For the enzymatic clean-up a minimum volume of 10 µl is required. To enable reproducable size selectiona minimum of 50 µl sample volume is required for the size selection protocols.
For the Right Side Size Selection protocol and the Double Sided Size Selection protocol only column 1 to 6 can be used for samples while using these protocols.
Sample positions for Left Side Size selection and Enzymatic clean-up protocol.
Sample positions for Right Side Size selection and Double Sided Size selection protocol.
D) Sample output position:
On the sample output position an empty plate must be placed. After the run this plate will contain thesample DNA. Please note that the well position of the samples will be the same as the well position of thesamples in the input position.
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 6
MagnaMedicsMove to Simplicity
Step 3. Run the protocol
When finished with the preperation of the deck, click 'Continue & update', followed bythe Start button
Your MagSi-NGSPREP protocol is now being performed. The estimated handling times are stated in the table below.
Time to complete protocol
96 samples 48 samples 32 samples 16 samples
4.1 Clean-up of enzymatic reaction < 65 min < 45 min < 35 min < 25 min4.2 Left Side Size Selection
4.3 Right Side Size Selection
N.A. < 50 min < 40 min < 30 min4.4 Double Sided Size Selection
When the protocol has been completed, the Primacontroller software displays a message, and the arm of the PrimaDiag platform moves to the homing position.
If you have any questions or require further help, please contact MagnaMedics at support@magnamedics.com for assistance to automate your individual laboratory workflow with MagSi-NGSPREP
Software or software protocols (e.g. for liquid handling systems) are provided under European Union Public Licence (EUPL v.1.1). The
Software may be changed or adopted under these regulation. The usage of the MagnaMedics software protocols is only permitted in
combination with the usage of Contractual Products.
The software protocols offered by MagnaMedics are provided "as is" and any expressed or implied warranties, including, but not limited
to, the implied warranties of merchantability and fitness for a particular purpose are disclaimed. In no event shall MagnaMedics, or any of
her contributors be liable for any direct, indirect, incidental, special, exemplary, or consequential damages (including, but not limited to,
procurement of substitute goods or services; loss of use, data, or profits; or business interruption) however caused and on any theory of
liability, whether in contract, strict liability, or tort (including negligence or otherwise) arising in any way out of the use of this software,
even if advised of the possibility of such damage.
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 7
MagnaMedicsMove to Simplicity
HowToAutomateMagSi-NGSPREPonPrimaDiag_v1-0_140429 8
MagnaMedicsMove to Simplicity
Recommended