New version of TWIST wf

[cesve]

Lims Issue

Work Flow: TWIST v5 Protocol: And here! Step: Here to!

Description:

Current set-up:

• [x] Change protocol Pre Processing TWIST v2

Steps: Reception Control TWIST (no change)

New step: Sample placement.

Process UDF's: Method document Document version Lot Nr: Nuclease-free water

Script for placement map (original well) Script for copying sample volume Copy Volume (uL) from Sample

• [x] Script for calculating volumes (see Calculate Volumes) PR for new EPP here. Merge to master at the same time as deploying the workflow

Script for creating Hamilton normalization file (probably already exists)

Step: Buffer Exchange TWIST Copy total volume from previous step, can this be added to UDF Sample volume (ul)?

• [x] Update script Calculate Volumes PR for new EPP here. Merge to master at the same time as deploying the workflow

• [x] Change protocol Target Enrichment TWIST v2

Steps: Pool samples for hybridization (no change)

Merge steps Hybridize Library TWIST, Capture and Wash TWIST, Amplify Captured Libraries TWIST and Bead Purification (post hybridization) TWIST into one step Target enrichment TWIST.

Process UDF's: Blockers TWIST Hybridization kit Thermal cycler (hyb) *new Hyb time start Method document Document version

Nuclease-free water Total hybridization time (h) Twist Wash Buffers Twist Binding and Purification beads

xGen Library Amp primer Kapa HiFi HotStart ReadyMix Thermal cycler (amplification) *new

Ethanol

Sample UDF's: Sample Name LIMS ID (Submitted Sample) Well Capture Library version Capture kit lot no. Bait Set Container Name Nr of PCR cycles *new PCR plate *new

Check: script for checking pcr-plate and Nr of cycles.

• [x] New protocol: Normalization of samples for sequencing (TWIST) Step: Normalization of samples for sequencing (TWIST)

• [x] New protocol: Library Validation QC TWIST (post norm) This should be an exact copy of Library Validation QC TWIST (post hyb) with the TapeStation step. Sizes are copied from previous QC-protocol. Set QC should be set manually.

[cesve]

New protocol: Pre Processing TWIST v2. Arnold and script for Skip buffer exchange are probably not affected by this change!

[cesve]

Changed protocol Target Enrichment TWIST v2:

Arnold needs to be changed in following models:

https://github.com/Clinical-Genomics/cg_lims/blob/master/cg_lims/models/arnold/prep/twist/hybridize_library_twist.py

https://github.com/Clinical-Genomics/cg_lims/blob/master/cg_lims/models/arnold/prep/twist/capture_and_wash_twist.py

https://github.com/Clinical-Genomics/cg_lims/blob/master/cg_lims/models/arnold/prep/twist/amplify_captured_libraries.py

https://github.com/Clinical-Genomics/cg_lims/blob/master/cg_lims/models/arnold/prep/twist/bead_purification_twist.py

Add new model for protocol "Normalization of samples for sequencing (TWIST)" Step: "Normalization of samples for sequencing (TWIST)"

[cesve]

• [x] Script Calculate Missing Reads For Pools. Move to Normalization of samples for sequencing (TWIST).
• [x] Script Arnold Upload twist . Move to Aggregate QC (Library Validation) TWIST (post norm).
• [x] Script Set Library QC to sample. Move to Aggregate QC (Library Validation) TWIST (post hyb). Already here?
• [x] Update script Copy Concentration (nM) from Library Preparation or Sample-Artifact.
• [ ] Update Script Copy Concentration (nM) from Library Preparation or Sample-Artifact when all samples from Twist v5 have been sequenced.

[cesve]

When deploying remember to:

• [x] Deactivate Copy Volume (ul) from Sample in step Buffer Exchange v2

[cesve]

_Test of new workflow Protocol: _Pre Processing TWIST v3__

Samples included in the test and expected values:

Step: Reception Control TWIST

Samples chosen for buffer exchange/skip buffer exchange. A barcode was given to the samples.

Step: Sample Placement (TWIST)

Script Copy Volume (ul) from Sample worked well and fetched the correct values.

Script for creating Placement Map worked well. Values are correct and it is placed at the correct placeholder.

Script Calculate Water calculated the correct volumes. When UDF Sample Volume (ul) is below 50, water is added to a total volume of 50 ul. The UDF Total Volume (uL) is Sample Volume (ul) + Volume H2O (ul). When the Sample Volume (ul) was above 100 ul a warning appeared.

Step: Buffer Exchange TWIST

Script Copy Sample Volume (ul) from Sample Placement (TWIST) copied the UDF Total Volume (uL) from previous step to UDF Sample Volume (ul) in this step. Values are correct.

Script Calculate Volumes calculated the correct Volume Beads (ul) (Sample Volume (ul) 2) and the correct Volume Elution (ul) (Sample Volume (ul) 0.8).

Script for creating Placement Map worked well and shown Source container and Source Well.

The correct Volumes (ul) are imported in Initial QC (DNA) TWIST protocol.

All steps and scripts in Protocol Pre Processing TWIST v3 worked as expected!

[cesve]

Note: Samples that are chosen to Skip Buffer Exchange are transferred to old workflow (TWIST v5).

Fixed!

[cesve]

Test of new workflow Protocol: Target Enrichment TWIST v3

Step: Pool samples for hybridization

No changes but to make the Hamilton Normalization File to work a Sample Placement view is needed. In addition the Placeholder name Placement Map was changed to Pooling Map to reflect what it actually is.

Volumes are correct.

Step: Target enrichment TWIST

When leaving fields empty a warning appears and it's not possible to continue to next step.

[cesve]

Test of new workflow Protocol: Normalization of samples for sequencing (TWIST)

Step: Normalization of samples for sequencing (TWIST)

The correct concentrations are imported. The Placement Map is correct.

[cesve]

Test of new workflow Protocol: Library Validation QC TWIST (post norm)

Step: Qubit QC (Library Validation) TWIST

Step: Quantit QC (Library Validation) TWIST

Step: Tapestation QC TWIST

[cesve]

Check to update automation configurations referring to steps in the old workflow:

Removed Mastersteps:

1. Hybridize Library TWIST v2
2. Capture and Wash TWIST v2
3. Amplify Captured Libraries TWIST v2
4. Bead Purification TWIST v2

Running check config:

(cg_lims) mayabrmbp:cg_lims maya.brandi$ lims -c ../config/genologics-stage.yaml scripts -l log check-config  --automation-string 'Hybridize Library TWIST v2'
(cg_lims) mayabrmbp:cg_lims maya.brandi$ lims -c ../config/genologics-stage.yaml scripts -l log check-config  --automation-string 'Capture and Wash TWIST v2'
(cg_lims) mayabrmbp:cg_lims maya.brandi$ lims -c ../config/genologics-stage.yaml scripts -l log check-config  --automation-string 'Amplify Captured Libraries TWIST v2'
(cg_lims) mayabrmbp:cg_lims maya.brandi$ lims -c ../config/genologics-stage.yaml scripts -l log check-config  --automation-string 'Bead Purification TWIST v2'

log output:

INFO:cg_lims.scripts.check_config:AUTOMATION: Searching for 'Hybridize Library TWIST v2'.

INFO:cg_lims.scripts.check_config:AUTOMATION: Searching for 'Capture and Wash TWIST v2'.

INFO:cg_lims.scripts.check_config:AUTOMATION: Searching for 'Amplify Captured Libraries TWIST v2'.

INFO:cg_lims.scripts.check_config:AUTOMATION: Searching for 'Bead Purification TWIST v2'.

INFO:cg_lims.scripts.check_config:Button: Copy Concentration (nM) from Library Preparation or Sample-Artifact
INFO:cg_lims.scripts.check_config:Bash string: bash -c -l "conda activate cg_lims && lims -c /home/glsai/.genologics.yaml epps -l {compoundOutputFileLuid0} -p {processLuid} udf copy artifact-to-artifact  -n 'Library Prep (Dev) v3' -n 'CG002 - RML Library Pooling' -n 'Normalization of RNA samples for sequencing v1' -n 'Pooling and Clean-up (Cov) v1' -n 'Bead Purification TWIST v2' -n 'End repair Size selection A-tailing and Adapter ligation (TruSeq PCR-free DNA)' -n  --sample-artifact -au  'Concentration (nM)'"
INFO:cg_lims.scripts.check_config:Processes: [('1006', 'Define Run Format and Calculate Volumes (Nova Seq)')]

• [x] Update Button Copy Concentration (nM) from Library Preparation or Sample-Artifact when deploying

[mayabrandi]

• [x] update arnold models
  • New protocol: Target Enrichment TWIST v3
  • New Protocol: Normalization of samples for sequencing (TWIST)
  • New protocol: Library Validation QC TWIST (post norm) values saved on normalization output

[cesve]

Test in LIMS production

Project #259917 ACC9643A1-A8 1508_25_Test220323.xlsx

Protocol: Pre Processing TWIST v3 Reception Control TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/311700 Sample Placement (TWIST): https://clinical-lims.scilifelab.se/clarity/work-details/311901 Buffer Exchange TWIST: https://clinical-lims.scilifelab.se/clarity/work-details/311902

Protocol: Initial QC TWIST v3 Tapestation Reception Control TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/311907 https://clinical-lims.scilifelab.se/clarity/work-complete/312103 Quantit QC (DNA) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/311908 Qubit QC (DNA) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312105 Aggregate QC (DNA) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/311909 https://clinical-lims.scilifelab.se/clarity/work-complete/312106

Protocol: Library Prep TWIST v2 Aliquot samples for enzymatic fragmentation TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312107 https://clinical-lims.scilifelab.se/clarity/work-details/311911 Enzymatic fragmentation TWIST: https://clinical-lims.scilifelab.se/clarity/work-details/311913 https://clinical-lims.scilifelab.se/clarity/work-details/312108 KAPA Library Preparation: https://clinical-lims.scilifelab.se/clarity/work-complete/311918 https://clinical-lims.scilifelab.se/clarity/work-complete/312109

Protocol: Library Validation QC TWIST v2 Tapestation QC TWIST: https://clinical-lims.scilifelab.se/clarity/work-details/311920 https://clinical-lims.scilifelab.se/clarity/work-complete/312110 Quantit QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-details/311924 Qubit QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312112 Aggregate QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-details/311926 https://clinical-lims.scilifelab.se/clarity/work-details/312113

Protocol: Target Enrichment TWIST v3 Pool samples for hybridization TWIST: https://clinical-lims.scilifelab.se/clarity/work-details/311929 https://clinical-lims.scilifelab.se/clarity/work-details/312121 Target enrichment TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/311930 https://clinical-lims.scilifelab.se/clarity/work-complete/312122

Protocol: Library Validation QC TWIST (post hyb) Tapestation QC TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/311931 https://clinical-lims.scilifelab.se/clarity/work-complete/312125 Quantit QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/311932 Qubit QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312126 Aggregate QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312418 https://clinical-lims.scilifelab.se/clarity/work-complete/312420

Protocol: Normalization of samples for sequencing (TWIST) v1 Normalization of samples for sequencing (TWIST): https://clinical-lims.scilifelab.se/clarity/work-details/312422

Protocol: Library Validation QC TWIST (post norm) Qubit QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312424 Quantit QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312426 Aggregate QC (Library Validation) TWIST: https://clinical-lims.scilifelab.se/clarity/work-complete/312425 https://clinical-lims.scilifelab.se/clarity/work-complete/312427

[cesve]

Due to human error the values in Normalization of samples for sequencing (TWIST): was accidentally overwritten manually. To test the script "Copy Concentration from Target Enrichment TWIST" one pool was re-queued to this step. The script copies the correct concentration.

https://clinical-lims.scilifelab.se/clarity/work-details/312702