Library Preparation v1.1

Protocol 1: Library Prep (AmpliSeq for Illumina Custom DNA Panel v1.1)

Protocol Type = Library Prep

Next Steps Configuration

Step 1: Dilute DNA (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Dilute DNA (AmpliSeq for Illumina Custom DNA Panel)
Step Type = Standard
Derived Sample Generation = Fixed, 1
Naming Convention = {InputItemName}
Reagent Kits
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Copy Concentration and Set Sample Volume

Trigger Location = Record Details
Trigger Style = Automatic upon entry

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Concentration:: = input.::Concentration:: ; output.::Conc. Units:: = input.::Conc. Units:: ; output.::Sample Volume (uL):: = 5' \
-log {compoundOutputFileLuid0}"

Calculate Input Amount, Total Volume & Low TE - Custom DNA Panel

Trigger Location = Record Details
Trigger Style = Manual button

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
        -t true \
        -h false \
        -exp 'output.::Input Amount (ng):: = output.::Concentration:: * output.::Sample Volume (uL):: ; \
output.::Total Volume (uL):: = output.::Input Amount (ng):: / step.::Desired Concentration (ng/uL):: ; \
output.::Low TE (uL):: = output.::Total Volume (uL):: - output.::Sample Volume (uL)::' \
        -log {compoundOutputFileLuid0}"

Calc Input Amount, Total Volume & Low TE - Custom DNA - Final Conc.

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
        -t true \
        -h false \
        -exp 'output.::Input Amount (ng):: = output.::Concentration:: * output.::Sample Volume (uL):: ; \
output.::Total Volume (uL):: = output.::Input Amount (ng):: / step.::Desired Final Concentration (ng/uL):: ; \
output.::Low TE (uL):: = output.::Total Volume (uL):: - output.::Sample Volume (uL):: ; \
output.::Desired Final Concentration (ng/uL):: = step.::Desired Final Concentration (ng/uL)::' \
        -log {compoundOutputFileLuid0}"

Set Next Step - Advance

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields

Step 2: Qubit (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Qubit (AmpliSeq for Illumina v1.1)
Step Type = Standard QC
Measurement Generation = Fixed, 1
Naming Convention = {InputItemName}

Automations

Assign QC flags (Qubit QC)

Trigger Location = Record Details
Trigger Style = Manual button

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {processURI:v2} -u {username} -p {password} script:assignQC -log {compoundOutputFileLuid1} -qcResult {compoundOutputFileLuid2}"

Set Next Step and Copy Concentration

Trigger Location = Record Details
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} script:evaluateDynamicExpression -t false -h false -exp 'nextStep = ::ADVANCE:: ; input.::Concentration:: = output.::Concentration:: ; input.::Conc. Units:: = output.::Conc. Units::' -log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
- QC Log File - Automatically attached
- QC Result File - Automatically attached
- Upload File - Manually uploaded
Sample Table
- Enable QC Flags = Yes
- Sample Display Default = Expand
- Well Sort Order = Row
- File Column Options
  - File Column Display = Hide
  - File Attachment Method = Auto
- Table Columns - Global Fields

Step 3: Dilute DNA to Final Concentration (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Dilute DNA (AmpliSeq for Illumina Custom DNA Panel)
Step Type = Standard
Derived Sample Generation = Fixed, 1
Naming Convention = {InputItemName}
Reagent Kits
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Copy Concentration and Set Sample Volume

Trigger Location = Record Details
Trigger Style = Automatic upon entry

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Concentration:: = input.::Concentration:: ; output.::Conc. Units:: = input.::Conc. Units:: ; output.::Sample Volume (uL):: = 5' \
-log {compoundOutputFileLuid0}"

Calc Input Amount, Total Volume & Low TE - Custom DNA - Final Conc.

Trigger Location = Record Details
Trigger Style = Manual button

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
        -t true \
        -h false \
        -exp 'output.::Input Amount (ng):: = output.::Concentration:: * output.::Sample Volume (uL):: ; \
output.::Total Volume (uL):: = output.::Input Amount (ng):: / step.::Desired Final Concentration (ng/uL):: ; \
output.::Low TE (uL):: = output.::Total Volume (uL):: - output.::Sample Volume (uL):: ; \
output.::Desired Final Concentration (ng/uL):: = step.::Desired Final Concentration (ng/uL)::' \
        -log {compoundOutputFileLuid0}"

Calculate Input Amount, Total Volume & Low TE - Custom DNA Panel

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
        -t true \
        -h false \
        -exp 'output.::Input Amount (ng):: = output.::Concentration:: * output.::Sample Volume (uL):: ; \
output.::Total Volume (uL):: = output.::Input Amount (ng):: / step.::Desired Concentration (ng/uL):: ; \
output.::Low TE (uL):: = output.::Total Volume (uL):: - output.::Sample Volume (uL)::' \
        -log {compoundOutputFileLuid0}"

Set Next Step - Advance

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields

Step 4: Amplify Targets - One Primer Pool (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Amplify Targets (AmpliSeq for Illumina Custom DNA Panel v1.1)
Step Type = Standard
Derived Sample Generation = Fixed, 1
Naming Convention = {InputItemName}
Reagent Kits
- AmpliSeq Custom DNA Panel for Illumina
  - Supplier = Illumina
  - Catalog Number = 20020495
  - Website = www.illumina.com
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Copy Desired Final Conc. & Calculate Master Mix, Total Sample, DNA Volume, Water - Custom DNA 1 Primer Pool

Trigger Location = Record Details
Trigger Style = Manual button

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::Master Mix (uL):: = 14 ; \
step.::Total Samples:: = step.::Total Samples:: + 1 ; \
step.::5X AmpliSeq HiFi Mix (uL):: = step.::Total Samples:: *1.1* 4 ; \
step.::2X AmpliSeq Custom DNA Panel:: = step.::Total Samples:: * 1.1 * 10 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 6) {output.::Nuclease-free water (uL):: = 20 -(output.::Master Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Set Next Step - Advance

Trigger Location = Record Details
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 2 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 7.5) {output.::Nuclease-free water (uL):: = 12.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 3 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 10.5) {output.::Nuclease-free water (uL):: = 17.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 4 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 9 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 13.5) {output.::Nuclease-free water (uL):: = 22.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Placement = Enabled

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
- Placement Pattern = Column
Destination Containers
- 96 well plate

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Column
- Table Columns - Global Fields

Step 5: Amplify Targets - Two Primer Pools (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Amplify Targets (AmpliSeq for Illumina Custom DNA Panel v1.1)
Step Type = Standard
Derived Sample Generation = Fixed, 1
Naming Convention = {InputItemName}
Reagent Kits
- AmpliSeq Custom DNA Panel for Illumina
  - Supplier = Illumina
  - Catalog Number = 20020495
  - Website = www.illumina.com
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 2 Primer Pool

Trigger Location = Record Details
Trigger Style = Manual button

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 7.5) {output.::Nuclease-free water (uL):: = 12.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Set Next Step - Advance

Trigger Location = Record Details
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Copy Desired Final Conc. & Calculate Master Mix, Total Sample, DNA Volume, Water - Custom DNA 1 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::Master Mix (uL):: = 14 ; \
step.::Total Samples:: = step.::Total Samples:: + 1 ; \
step.::5X AmpliSeq HiFi Mix (uL):: = step.::Total Samples:: *1.1* 4 ; \
step.::2X AmpliSeq Custom DNA Panel:: = step.::Total Samples:: * 1.1 * 10 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 6) {output.::Nuclease-free water (uL):: = 20 -(output.::Master Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 3 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 10.5) {output.::Nuclease-free water (uL):: = 17.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 4 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 9 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 13.5) {output.::Nuclease-free water (uL):: = 22.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Placement = Enabled

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
- Placement Pattern = Column
Destination Containers
- 96 well plate

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields

Step 6: Amplify Targets - Three Primer Pool (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Amplify Targets (AmpliSeq for Illumina Custom DNA Panel v1.1)
Step Type = Standard
Derived Sample Generation = Fixed, 1
Naming Convention = {InputItemName}
Reagent Kits
- AmpliSeq Custom DNA Panel for Illumina
  - Supplier = Illumina
  - Catalog Number = 20020495
  - Website = www.illumina.com
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 3 Primer Pool

Trigger Location = Record Details
Trigger Style = Manual button

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 10.5) {output.::Nuclease-free water (uL):: = 17.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Set Next Step - Advance

Trigger Location = Record Details
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Copy Desired Final Conc. & Calculate Master Mix, Total Sample, DNA Volume, Water - Custom DNA 1 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::Master Mix (uL):: = 14 ; \
step.::Total Samples:: = step.::Total Samples:: + 1 ; \
step.::5X AmpliSeq HiFi Mix (uL):: = step.::Total Samples:: *1.1* 4 ; \
step.::2X AmpliSeq Custom DNA Panel:: = step.::Total Samples:: * 1.1 * 10 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 6) {output.::Nuclease-free water (uL):: = 20 -(output.::Master Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 2 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 7.5) {output.::Nuclease-free water (uL):: = 12.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 4 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 9 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 13.5) {output.::Nuclease-free water (uL):: = 22.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields

Step 7: Amplify Targets - Four Primer Pool (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Amplify Targets (AmpliSeq for Illumina Custom DNA Panel v1.1)
Step Type = Standard
Derived Sample Generation = Fixed, 1
Naming Convention = {InputItemName}
Reagent Kits
- AmpliSeq Custom DNA Panel for Illumina
  - Supplier = Illumina
  - Catalog Number = 20020495
  - Website = www.illumina.com
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 4 Primer Pool

Trigger Location = Record Details
Trigger Style = Manual button

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 9 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 13.5) {output.::Nuclease-free water (uL):: = 22.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Set Next Step - Advance

Trigger Location = Record Details
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Copy Desired Final Conc. & Calculate Master Mix, Total Sample, DNA Volume, Water - Custom DNA 1 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::Master Mix (uL):: = 14 ; \
step.::Total Samples:: = step.::Total Samples:: + 1 ; \
step.::5X AmpliSeq HiFi Mix (uL):: = step.::Total Samples:: *1.1* 4 ; \
step.::2X AmpliSeq Custom DNA Panel:: = step.::Total Samples:: * 1.1 * 10 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 6) {output.::Nuclease-free water (uL):: = 20 -(output.::Master Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 2 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 7.5) {output.::Nuclease-free water (uL):: = 12.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Copy Desired DNA Conc. and Calc DNA Vol & Water - Custom DNA 3 Primer Pool

Trigger Location = Not Used

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
script:evaluateDynamicExpression \
-t false \
-h false \
-exp 'output.::Diluted DNA Conc. (ng/uL):: = input.::Desired Final Concentration (ng/uL):: ; \
output.::5X AmpliSeq HiFi Mix (uL):: = 5 ; \
output.::DNA Volume (uL):: = output.::Prep Input Amount (ng):: / output.::Diluted DNA Conc. (ng/uL):: ; \
if (output.::DNA Volume (uL):: < 10.5) {output.::Nuclease-free water (uL):: = 17.5 - (output.::5X AmpliSeq HiFi Mix (uL):: + output.::DNA Volume (uL)::)}' \
-log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Placement = Enabled

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
- Placement Pattern = Column
Destination Containers
- 96 well plate

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields

Step 8: Partially Digested Amplicons (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Partially Digested Amplicons (AmpliSeq for Illumina v1.1)
Step Type = No Outputs
Reagent Kits
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Set Next Step - Advance

Trigger Location = Record Details
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Queue/Ice Bucket

Defaults
- Sample Grouping = Group by Containers
- Well Sort Order = Row
Sample Table (Column Headers)

Record Details

Step Data (Master Step Fields)
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields

Step 9: Ligate Indexes (AmpliSeq for Illumina Custom DNA Panel v1.1)

Master Step Name = Ligate Indexes (AmpliSeq for Illumina v1.1)
Step Type = Add Labels
Derived Sample Generation = Fixed, 1
Naming Convention = {InputItemName}
Reagent Kits
- AmpliSeq Library PLUS for Illumina
  - Supplier = Illumina
  - Catalog Number = 24 - 20019101; 96 - 200191102; 384 - 200191103
  - Website = www.illumina.com

Automations

Set Next Step - Advance

Trigger Location = Record Details
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} \
      script:evaluateDynamicExpression \
      -t false \
      -h false \
      -exp 'nextStep = ::ADVANCE::' \
      -log {compoundOutputFileLuid0}"

Route AmpliSeq Samples - Custom DNA Panel

Trigger Location = Step
Trigger Style = Automatic upon exit

bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -u {username} -p {password} -i {stepURI:v2} -l {compoundOutputFileLuid0} script:changeWorkflow \
\
--FIELD_NAME 'AmpliSeq Workflow' \
--FIELD_VALUE 'Standard Workflow' \
--WORKFLOW 'Standard Workflow AmpliSeq for Illumina Custom DNA Panel v1.1' \
--STEP 'Clean Up Library (AmpliSeq for Illumina Custom DNA Panel v1.1)' \
--INPUTS_OR_OUTPUTS 'OUTPUTS' \
\
--FIELD_NAME 'AmpliSeq Workflow' \
--FIELD_VALUE 'Equalizer Workflow' \
--WORKFLOW 'Equalizer Workflow AmpliSeq for Illumina Custom DNA Panel v1.1' \
--STEP 'Clean Up Library (AmpliSeq for Illumina Custom DNA Panel v1.1)' \
--INPUTS_OR_OUTPUTS 'OUTPUTS'"