TruSeq RNA Access v2.0

Overview

The TruSeq RNA Access v2.0 workflow includes the following functionality:

• Preconfigured TruSeq RNA Access v2.0 protocol that supports the conversion of total RNA or FFPE samples into a library of template molecules suitable for subsequent cluster generation and sequencing.
• Automated calculation of sample and buffer volumes.
• Automated calculation or display of reagents at every step in the workflow.
• Automatic step transition when required.
• Automatic placement of samples when necessary.
• Automated generation of Bioanalyzer input file that the user may upload to the Bioanalyzer instrument’s Expert Software and use to set up a run.
• Automated parsing of a Bioanalyzer instrument's XML result file into the LIMS, and storing of QC measurement records for the completed run.
• Automated assignment of QC Pass/Fail, based on user-selected threshold values.
• Routing script that allows sequencing of libraries using any Illumina sequencing instrument.

Protocol 1: TruSeq RNA Access v2.0

Protocol Type = Library Prep

Next Steps Configuration

Step 1: Sort Samples (TruSeq RNA Access v2.0)

• Master Step Name = Sort Samples (TruSeq RNA Access v2.0.10)
• Step Type = No Outputs

The version of Sort Samples master step name may be different depending on the version of IPP installed.

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	Well	Built-in
    Derived Sample	Sample Name	Built-in
    Derived Sample	Waiting	Built-in

  • Expanded View Fields

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

Record Details

• Step Data (Master Step Fields)

  Field Name	Field Type	Options	Additional Options and Dropdown Items
  Comment	Multiline Text

• Sample Table

  • Sample Display Default = Collapse
  • 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 2: Fragment RNA (TruSeq RNA Access v2.0)

• Master Step Name = Fragment RNA v2.0
• Step Type = Standard
• Derived Sample Generation = Fixed, 1
• Naming Convention = {SubmittedSampleName}
• Reagent Kits

  • TruSeq RNA Access - 12 Index Set A or B, Box 2

    • Supplier = Illumina
    • Catalog Number = Set A Part # 15052311; Set B Part # 15052312
    • Website = https://support.illumina.com/

Automations

Dilute Total RNA

• Trigger Location = Record Details
• Trigger Style = Manual button

Set Next Step - Advance

• Trigger Location = Record Details
• Trigger Style = Automatic upon exit

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	Well	Built-in
    Derived Sample	Sample Name	Built-in
    Derived Sample	Waiting	Built-in

  • Expanded View Fields

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

Placement

• Defaults

  • Sample Grouping = Group by Containers
  • Well Sort Order = Row
  • Placement Pattern = Row
• Destination Containers

  • 96 well plate

Record Details

• Step Data (Master Step Fields)

  Field Name	Field Type	Options	Additional Options and Dropdown Items
  Comment	Multiline Text
  Thermal Cycler Program	Text		Default = Elution 2 - Frag - Prime

• Step File Placeholders

  • Log File - 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	Buffer Volume (ul)	Numeric		Decimal Places Displayed = 2
    Derived Sample	Sample Name	Built-in
    Derived Sample	Sample Volume (ul)	Numeric		Decimal Places Displayed = 2
    Project	Project Name	Built-in

Step 3: First Strand cDNA Synthesis (TruSeq RNA Access v2.0)

• Master Step Name = First Strand cDNA Synthesis v2.0
• Step Type = No Outputs
• Reagent Kits

  • TruSeq RNA Access - cDNA Synthesis-PCR, Box 1

    • Supplier = Illumina
    • Catalog Number = 15052309
    • Website = https://support.illumina.com/

Automations

Set Next Step - Advance

• Trigger Location = Record Details
• Trigger Style = Automatic upon exit

Queue/Ice Bucket

• Defaults

  • Sample Grouping = Group by Containers
  • Well Sort Order = Column
• Sample Table

  • Column Headers

    Category	Field Name	Field Type	Options	Additional Options and Dropdown Items
    Container	Container Name	Built-in
    Container	Well	Built-in
    Derived Sample	Sample Name	Built-in
    Derived Sample	Waiting	Built-in

  • Expanded View Fields

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

Record Details

• Step Data (Master Step Fields)

  Field Name	Field Type	Options	Additional Options and Dropdown Items
  Comment	Multiline Text
  Thermal Cycler Program	Text Dropdown	Custom Entries	Presets Synthesize 1st Strand 1stSS Default = Synthesize 1st Strand

• Step File Placeholders

  • Log File - 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 4: Second Strand cDNA Synthesis (TruSeq RNA Access v2.0)

• Master Step Name = Second Strand cDNA Synthesis v2.0
• Step Type = Standard
• Derived Sample Generation = Fixed, 1
• Naming Convention = {SubmittedSampleName}
• Reagent Kits

  • TruSeq RNA Access - cDNA Synthesis-PCR, Box 1

    • Supplier = Illumina
    • Catalog Number = 15052309
    • Website = https://support.illumina.com/

Automations

Set Next Step - Advance

• Trigger Location = Record Details
• Trigger Style = Automatic upon exit

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	Well	Built-in
    Derived Sample	Sample Name	Built-in
    Derived Sample	Waiting	Built-in

  • Expanded View Fields

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

Record Details

• Step Data (Master Step Fields)

  Field Name	Field Type	Options	Additional Options and Dropdown Items
  Comment	Multiline Text
  80% EtOH Prep Date	Date

• Step File Placeholders

  • Log File - 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	Sample Name	Built-in
    Project	Project Name	Built-in

Step 5: Adenylate 3' Ends (TruSeq RNA Access v2.0)

• Master Step Name = Adenylate 3' Ends v2.0
• Step Type = No Outputs
• Reagent Kits

  • TruSeq RNA Access - 12 Index Set A or B, Box 2

    • Supplier = Illumina
    • Catalog Number = Set A Part # 15052311; Set B Part # 15052312
    • Website = https://support.illumina.com/

Automations

Set Next Step - Advance

• Trigger Location = Record Details
• Trigger Style = Automatic upon exit

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	Well	Built-in
    Derived Sample	Sample Name	Built-in
    Derived Sample	Waiting	Built-in

  • Expanded View Fields

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

Record Details

• Step Data (Master Step Fields)

  Field Name	Field Type	Options	Additional Options and Dropdown Items
  Comment	Multiline Text

• Step File Placeholders

  • Log File - 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	Sample Name	Built-in
    Project	Project Name	Built-in

Step 6: Ligate Adapters (TruSeq RNA Access v2.0)

• Master Step Name = Ligate Adapters v2.0
• Step Type = Add Labels
• Derived Sample Generation = Fixed, 1
• Naming Convention = {SubmittedSampleName}
• Reagent Kits

  • TruSeq RNA Access - 12 Index Set A or B, Box 2

    • Supplier = Illumina
    • Catalog Number = Set A Part # 15052311; Set B Part # 15052312
    • Website = https://support.illumina.com/

Automations

Set Next Step - Advance

• Trigger Location = Record Details
• Trigger Style = Manual button

Copy to Output

• Trigger Location = Not Used

Set Next Step & Copy to Input

• Trigger Location = Not Used

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	Well	Built-in
    Derived Sample	Sample Name	Built-in
    Derived Sample	Waiting	Built-in

  • Expanded View Fields

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

Add Labels

• Label Groups

  • TruSeq RNA Access

Record Details

• Step Data (Master Step Fields)

  Field Name	Field Type	Options	Additional Options and Dropdown Items
  Comment	Multiline Text
  80% EtOH Prep Date	Date

• Step File Placeholders

  • Log File - 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	Reagent Name	Built-in
    Derived Sample	Sample Name	Built-in
    Project	Project Name	Built-in

Step 7: First PCR Amplification (TruSeq RNA Access v2.0)

• Master Step Name = First PCR Amplification (TruSeq RNA Access v2.0.10)
• Step Type = Standard
• Derived Sample Generation = Fixed, 1
• Naming Convention = {SubmittedSampleName}
• Reagent Kits

  • TruSeq RNA Access - cDNA Synthesis-PCR, Box 1

    • Supplier = Illumina
    • Catalog Number = 15052309
    • Website = https://support.illumina.com/

The version of First PCR Amplification master step name may be different depending on the version of IPP installed.

Automations

Set Next Step - Advance

• Trigger Location = Record Details
• Trigger Style = Automatic upon exit

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	Well	Built-in
    Derived Sample	Sample Name	Built-in
    Derived Sample	Waiting	Built-in

  • Expanded View Fields

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

Record Details

• Step Data (Master Step Fields)

  Field Name	Field Type	Options	Additional Options and Dropdown Items
  Comment	Multiline Text
  80% EtOH Prep Date	Date
  Thermal Cycler Program	Text		Default = PCR

• Step File Placeholders

  • Log File - 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 8: Bioanalyzer QC (Library Validation) (TruSeq RNA Access v2.0)

• Master Step Name = Bioanalyzer QC (Library Validation) v2.0
• Step Type = Standard QC
• Measurement Generation = Fixed, 1
• Naming Convention = {InputItemName} Bioanalyzer

Automations

Generate Bioanalyzer Input file

• Trigger Location = Record Details
• Trigger Style = Automatic upon entry

Parse Bioanalyzer XML and assign QC flags

• Trigger Location = Record Details
• Trigger Style = Manual button

Parse Bioanalyzer XML, Assign QC flags, and Copy Concentrations

• Trigger Location = Record Details
• Trigger Style = Manual button

Set Next Step - Output PASS/FAIL

• Trigger Location = Record Details