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Custom DNA Panel

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

Record Details

Step Data (Master Step Fields)
Field Name
Field Type
Options
Additional Options and Dropdown Items
Desired Concentration (ng/uL)
Numeric
Range = 20 To 50
Directions
Multiline Text
Read Only
Default = If enough DNA is available, dilute to intermediate concentration of ~20-50 ng/uL.
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  Concentration
  Numeric
  Required Field
  Decimal Places Displayed = 2
  Derived Sample
  Conc. Units
  Numeric
  Required Field
  Derived Sample
  Input Amount (ng)
  Numeric
  Decimal Places Displayed = 0
  Derived Sample
  Low TE (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Sample Name
  Built-in
  Derived Sample
  Sample Volume (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Total Volume (uL)
  Numeric
  Decimal Places Displayed = 2
  Project
  Project Name
  Built-in

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

Record Details

Step Data (Master Step Fields)
Field Name
Field Type
Options
Additional Options and Dropdown Items
Criteria 1 - Operator
Text
Default = >=
Criteria 1 - Source Data Field
Text
Default = Concentration
Criteria 1 - Threshold Value
Numeric
Criteria 2 - Operator
Text
Default = <=
Criteria 2 - Source Data Field
Text
Default = Concentration
Criteria 2 - Threshold Value
Numeric
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
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  Concentration
  Numeric
  Required Field
  Decimal Places Displayed = 2
  Derived Sample
  Conc. Units
  Text
  Required Field
  Derived Sample
  Sample Name
  Built-in
  Measurement
  Concentration
  Numeric
  Decimal Places Displayed = 2
  Measurement
  Conc. Units
  Text
  Project
  Project Name
  Built-in

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

Record Details

Step Data (Master Step Fields)
Field Name
Field Type
Options
Additional Options and Dropdown Items
Desired Final Concentration (ng/uL)
Numeric
Range = 2 To 8
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  Concentration
  Numeric
  Required Field
  Decimal Places Displayed = 2
  Derived Sample
  Conc. Units
  Numeric
  Required Field
  Derived Sample
  Desired Final Concentration (ng/uL)
  Numeric
  Derived Sample
  Low TE (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Sample Name
  Built-in
  Derived Sample
  Sample Volume (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Total Volume (uL)
  Numeric
  Decimal Places Displayed = 2
  Project
  Project Name
  Built-in

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

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)
Field Name
Field Type
Options
Additional Options and Dropdown Items
2X AmpliSeq Custom DNA Panel
Numeric
Decimal Places Displayed = 1
5X AmpliSeq HiFi Mix (uL)
Numeric
Decimal Places Displayed = 1
PCR Cycles
Numeric
Recommended Prep Input (ng)
Multiline Text
Read Only
Default = This protocol supports 1-100 ng per pool. The recommended input is 10 ng high-quality DNA per pool.
Thermal Cycler Program
Text
Default = AMP_DNA
Total Samples
Numeric
Default = 0
Decimal Places Displayed = 0
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Column
- Table Columns - Global Fields
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  Diluted DNA Conc. (ng/uL)
  Numeric
  Derived Sample
  DNA Volume (uL)
  Numeric
  Derived Sample
  Master Mix (uL)
  Numeric
  Derived Sample
  Nuclease-free water (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Prep Input Amount (ng)
  Numeric
  Required Field
  Range = 2 To 200
  Decimal Places Displayed = 0
  Derived Sample
  Sample Name
  Built-in
  Project
  Project Name
  Built-in

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

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)
Field Name
Field Type
Options
Additional Options and Dropdown Items
PCR Cycles
Numeric
Recommended Prep Input (ng)
Multiline Text
Read Only
Default = This protocol supports 1-100 ng per pool. The recommended input is 10 ng high-quality DNA per pool.
Thermal Cycler Program
Text
Default = AMP_DNA
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  5X AmpliSeq HiFi Mix (uL)
  Numeric
  Decimal Places Displayed = 1
  Derived Sample
  Diluted DNA Conc. (ng/uL)
  Numeric
  Derived Sample
  DNA Volume (uL)
  Numeric
  Derived Sample
  Nuclease-free water (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Prep Input Amount (ng)
  Numeric
  Required Field
  Range = 2 To 200
  Decimal Places Displayed = 0
  Derived Sample
  Sample Name
  Built-in
  Project
  Project Name
  Built-in

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

Record Details

Step Data (Master Step Fields)
Field Name
Field Type
Options
Additional Options and Dropdown Items
PCR Cycles
Numeric
Recommended Prep Input (ng)
Multiline Text
Read Only
Default = This protocol supports 1-100 ng per pool. The recommended input is 10 ng high-quality DNA per pool.
Thermal Cycler Program
Text
Default = AMP_DNA
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  5X AmpliSeq HiFi Mix (uL)
  Numeric
  Decimal Places Displayed = 1
  Derived Sample
  Diluted DNA Conc. (ng/uL)
  Numeric
  Derived Sample
  DNA Volume (uL)
  Numeric
  Derived Sample
  Nuclease-free water (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Prep Input Amount (ng)
  Numeric
  Required Field
  Range = 2 To 200
  Decimal Places Displayed = 0
  Derived Sample
  Sample Name
  Built-in
  Project
  Project Name
  Built-in

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

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)
Field Name
Field Type
Options
Additional Options and Dropdown Items
PCR Cycles
Numeric
Recommended Prep Input (ng)
Multiline Text
Read Only
Default = This protocol supports 1-100 ng per pool. The recommended input is 10 ng high-quality DNA per pool.
Thermal Cycler Program
Text
Default = AMP_DNA
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  5X AmpliSeq HiFi Mix (uL)
  Numeric
  Decimal Places Displayed = 1
  Derived Sample
  Diluted DNA Conc. (ng/uL)
  Numeric
  Derived Sample
  DNA Volume (uL)
  Numeric
  Derived Sample
  Nuclease-free water (uL)
  Numeric
  Decimal Places Displayed = 2
  Derived Sample
  Prep Input Amount (ng)
  Numeric
  Required Field
  Range = 2 To 200
  Decimal Places Displayed = 0
  Derived Sample
  Sample Name
  Built-in
  Project
  Project Name
  Built-in

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)
Category
Field Name
Field Type
Options
Additional Options and Dropdown Items
Container
Container Name
Built-in
Container
LIMS ID (Container)
Built-in
Container
Well
Built-in
Derived Sample
Sample Name
Built-in
Derived Sample
Waiting
Built-in
Project
Project Name
Built-in

Record Details

Step Data (Master Step Fields)
Field Name
Field Type
Options
Additional Options and Dropdown Items
Thermal Cycler Program
Text
Default = FUPA
Step File Placeholders
- Log - Automatically attached
Sample Table
- Sample Display Default = Expand
- Well Sort Order = Row
- Table Columns - Global Fields
  Category
  Field Name
  Field Type
  Options
  Additional Options and Dropdown Items
  Container
  Container Name
  Built-in
  Container
  LIMS ID (Container)
  Built-in
  Container
  Well
  Built-in
  Derived Sample
  Sample Name
  Built-in
  Project
  Project Name
  Built-in

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'"