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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.



http://www.magnamedics.com/software/primadiag/571-magsi-ngsprep

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


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.


4.3 Right SideSize Selection


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.


4.4 DoubleSided SizeSelection


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.




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



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


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




4.3 Right SideSize Selection

MagSi-NGSPREP 1 (Sample volume (µl) * 1.8) * Number of samples + 200 µl



4.4 DoubleSided SizeSelection

MagSi-NGSPREP 1(Sample volume (µl) * Left Ratio) * 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



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.



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 [email protected] 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.



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