# Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1

## Protocol 1: Prep Sample (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

Protocol Type = Sample Prep

**Next Steps Configuration**

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-4bff47fc6025ece89c208dab74022694f6a502f8%2Frna-ribo-protocol1-next-step-config.png?alt=media" alt=""><figcaption></figcaption></figure>

### Step 1: Hybridize Probes (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Hybridize Probes for Stranded Total RNA Prep v1.0
* Step Type = Standard
* Derived Sample Generation = Fixed, 1
* Naming Convention = {InputItemName}
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-1ae8ff01a9015dcea70edaec68d43a0be541a39f%2Frna-ribo-protocol1-step1-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Calculate Hybridize Probe Master Mix</summary>

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

{% code overflow="wrap" %}

```markup
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2:http} -u {username} -p {password} script:evaluateDynamicExpression -t false -h false \
-exp 'step.::Total Samples:: = step.::Total Samples:: + 1 ; step.::DB1 (uL):: = 3.6 * step.::Total Samples:: ; step.::DP1 (uL):: = 1.2 * step.::Total Samples::' \
-log {compoundOutputFileLuid0}"
```

{% endcode %}

</details>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                                                                                                     |
  | ---------------------------- | -------------- | ----------- | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
  | DB1 (uL)                     | Numeric        |             | <ul><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                                                |
  | DP1 (uL)                     | Numeric        |             | <ul><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                                                |
  | Instruction Notes            | Multiline Text | Read Only   | <ul><li>Default = Add 4 µl Hybridize Probe Master Mix to each well.</li></ul>                                                                                                                                 |
  | Thermal Cycler Program       | Text           |             | <ul><li>Default = HYB\_DP1</li></ul>                                                                                                                                                                          |
  | Thermal Cycler Program Notes | Multiline Text | Read Only   | <ul><li>Default =<br>Choose the preheat lid option and set to 100°C<br>Reaction volume is 15 µl u 95° for 2 minutes<br>Decrease 0.1°C per second until the temperature reaches 37°C<br>Hold at 37°C</li></ul> |
  | Total Samples                | Numeric        |             | <ul><li>Default = 0</li><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                            |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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 2: Deplete rRNA (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Deplete rRNA v1.0
* Step Type = No Outputs
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-465ad21de1e8c7c13ff5f02dea13dba75d679d91%2Frna-ribo-protocol1-step2-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Calculate rRNA Depletion Master Mix</summary>

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

{% code overflow="wrap" %}

```markup
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2:http} -u {username} -p {password} script:evaluateDynamicExpression -t false -h false \
-exp 'step.::Total Samples:: = step.::Total Samples:: + 1 ; step.::RDB (uL):: = 4.8 * step.::Total Samples:: ; step.::RDE (uL):: = 1.2 * step.::Total Samples::' \
-log {compoundOutputFileLuid0}"
```

{% endcode %}

</details>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                                                                              |
  | ---------------------------- | -------------- | ----------- | -------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
  | Instruction Notes            | Multiline Text | Read Only   | <ul><li>Default = Add 5 µl rRNA Depletion Master Mix to each well.</li></ul>                                                                                                           |
  | RDB (uL)                     | Numeric        |             | <ul><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                         |
  | RDE (uL)                     | Numeric        |             | <ul><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                         |
  | Thermal Cycler Program       | Text           |             | <ul><li>Default = RNA\_DEP</li></ul>                                                                                                                                                   |
  | Thermal Cycler Program Notes | Multiline Text | Read Only   | <ul><li>Default =<br>RNA\_DEP program on the thermal cycler:Choose the preheat lid option and set to 100°C<br>Reaction volume is 20 µl<br>37°C for 15 minutes<br>Hold at 4°C</li></ul> |
  | Total Samples                | Numeric        |             | <ul><li>Default = 0</li><li>Decimal Places Displayed = 0</li></ul>                                                                                                                     |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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 3: Remove Probes (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Remove Probes v1.0
* Step Type = No Outputs
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-342444dbf9c0b9d4396f7c4a2d9363151d4bec4e%2Frna-ribo-protocol1-step3-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Calculate Removal Master Mix</summary>

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

{% code overflow="wrap" %}

```markup
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2:http} -u {username} -p {password} script:evaluateDynamicExpression -t false -h false \
-exp 'step.::Total Samples:: = step.::Total Samples:: + 1 ; step.::PRB (uL):: = 7.7 * step.::Total Samples:: ; step.::PRE (uL):: = 3.3 * step.::Total Samples::' \
-log {compoundOutputFileLuid0}"
```

{% endcode %}

</details>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                                                                                                         |
  | ---------------------------- | -------------- | ----------- | ----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
  | Instruction Notes            | Multiline Text | Read Only   | <ul><li>Default = Add 10 µl Probe Removal Master Mix to each well.</li></ul>                                                                                                                                      |
  | PRB (uL)                     | Numeric        |             | <ul><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                                                    |
  | PRE (uL)                     | Numeric        |             | <ul><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                                                    |
  | Thermal Cycler Program       | Text           |             | <ul><li>Default = PRB\_REM</li></ul>                                                                                                                                                                              |
  | Thermal Cycler Program Notes | Multiline Text | Read Only   | <ul><li>Default =<br>PRB\_REM program on the thermal cycler:<br>Choose the preheat lid option and set to 100°C<br>Reaction volume is 30 µl<br>37°C for 15 minutes<br>70°C for 15 minutes<br>Hold at 4°C</li></ul> |
  | Total Samples                | Numeric        |             | <ul><li>Default = 0</li><li>Decimal Places Displayed = 0</li></ul>                                                                                                                                                |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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 4: Clean Up RNA (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Clean Up RNA v1.0
* Step Type = Standard
* Derived Sample Generation = Fixed, 1
* Naming Convention = {InputItemName}
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-ebedeae5b95ca2a1982b628762f6d872f3bea9e9%2Frna-ribo-protocol1-step4-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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 File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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: Fragment and Denature RNA (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Fragment and Denature RNA v1.0
* Step Type = No Outputs
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-e5919d17ca9ed8b01d1df633fd52bc4d688a64a6%2Frna-ribo-protocol1-step5-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                           |
  | ---------------------------- | -------------- | ----------- | ----------------------------------------------------------------------------------------------------------------------------------- |
  | Instruction Notes            | Multiline Text | Read Only   | Default = Add 8.5 µl EPH3 to each well.                                                                                             |
  | Thermal Cycler Program       | Text           |             | Default = DEN\_RNA                                                                                                                  |
  | Thermal Cycler Program Notes | Multiline Text | Read Only   | <p>Default =<br>Choose the preheat lid option and set to 100°C<br>Reaction volume is 17 µl<br>94°C for 2 minutes<br>Hold at 4°C</p> |
* Step File Placeholders
  * Log - Automatically attached
* 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 6: Synthesize First Strand cDNA (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Synthesize First Strand cDNA v1.0
* Step Type = No Outputs
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-6d98e0f1ffd6bff92897ee2d135c61f48d8d9aca%2Frna-ribo-protocol1-step6-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Calculate First Strand Synthesis Master Mix</summary>

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

{% code overflow="wrap" %}

```markup
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2:http} -u {username} -p {password} script:evaluateDynamicExpression -t false -h false -exp 'step.::Total Samples:: = step.::Total Samples:: + 1 ; step.::FSA (uL):: = 9 * step.::Total Samples:: ; step.::RVT (uL):: = 1 * step.::Total Samples:: ' -log {compoundOutputFileLuid0}"
```

{% endcode %}

</details>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                                                                          |
  | ---------------------------- | -------------- | ----------- | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
  | FSA (uL)                     | Numeric        |             | Decimal Places Displayed = 0                                                                                                                                                       |
  | Instruction Notes            | Multiline Text | Read Only   | Default = Add 8 uL of First Strand Synthesis Master Mix to each sample.                                                                                                            |
  | RVT (uL)                     | Numeric        |             | Decimal Places Displayed = 0                                                                                                                                                       |
  | Thermal Cycler Program       | Text           |             | Default = FSS                                                                                                                                                                      |
  | Thermal Cycler Program Notes | Multiline Text | Read Only   | <p>Default =<br>Choose the preheat lid option and set to 100°C<br>Reaction volume is 25 µl<br>25°C for 10 minutes<br>42°C for 15 minutes<br>70°C for 15 minutes<br>Hold at 4°C</p> |
  | Total Samples                | Numeric        |             | Default = 0                                                                                                                                                                        |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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 7: Synthesize Second Strand cDNA (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Synthesize Second Strand cDNA v1.0
* Step Type = Standard
* Derived Sample Generation = Fixed, 1
* Naming Convention = {InputItemName}
* Reagent Kits
  * Agencourt AMPure XP, 5 ml
    * Supplier = Beckman Coulter
    * Catalog Number = A63880
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-f8bcc77c7a37d5c42259003fc2f2f751301329de%2Frna-ribo-protocol1-step7-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                      |
  | ---------------------------- | -------------- | ----------- | ------------------------------------------------------------------------------------------------------------------------------ |
  | 80% EtOH Prep Date           | Date           |             |                                                                                                                                |
  | Thermal Cycler Program       | Text           |             | Default = SSS                                                                                                                  |
  | Thermal Cycler Program Notes | Multiline Text | Read Only   | <p>Default =<br>Choose the preheat lid option and set to 40°C<br>Reaction volume is 50 µl u 16°C for 1 hour<br>Hold at 4°C</p> |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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       |             |                                           |

## Protocol 2: Library Prep (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

Protocol Type = Library Prep

**Next Steps Configuration**

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-b64aea22eb5388826b3e466cc5d11e5844a225e3%2Frna-ribo-protocol2-next-step-config.png?alt=media" alt=""><figcaption></figcaption></figure>

### Step 1: Adenylate 3ʹ Ends (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Adenylate 3ʹ Ends - RNA Prep v1.0
* Step Type = No Outputs
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-4313e42da496e5dc8962571fc7062599f23b52eb%2Frna-ribo-protocol2-step1-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                                                  |
  | ---------------------------- | -------------- | ----------- | ---------------------------------------------------------------------------------------------------------------------------------------------------------- |
  | Instruction Notes            | Multiline Text | Read Only   | Default = Add 12.5 µl ATL4 to each well.                                                                                                                   |
  | Thermal Cycler Program       | Text           |             | Default = ATAIL                                                                                                                                            |
  | Thermal Cycler Program Notes | Multiline Text | Read Only   | <p>Default =<br>Choose the preheat lid option and set to 100°C<br>Reaction volume is 30 µl<br>37°C for 30 minutes<br>70°C for 5 minutes<br>Hold at 4°C</p> |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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 2: Ligate Anchors (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Ligate Anchors - RNA Prep v1.0
* Step Type = No Outputs
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-04742381bf51f008c5eff5bf9967afeb22f5d18a%2Frna-ribo-protocol2-step2-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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**                                                                                                                                                                                                                                   |
  | ----------------------------- | -------------- | ----------- | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
  | Add Anchors Instruction Notes | Multiline Text |             | <p>Default =<br>Volume for Sample Input <= 100 ng (µl)<br>1. Add 2.5 (µl) RSB<br>2. Add 2.5 (µl) RNA Index Anchors<br>3. Add 2.5 (µl) LIGX<br><br>Volume for Sample Input > 100 ng (µl)<br>1. Add 0 (µl) RSB<br>2. Add 5 (µl) RNA Index Anchors<br>3. Add 2.5 (µl) LIGX</p> |
  | Instruction Notes             | Multiline Text | Read Only   | Default = Add 5 µl STL to each well.                                                                                                                                                                                                                                        |
  | Thermal Cycler Program        | Text           |             | Default = LIG                                                                                                                                                                                                                                                               |
  | Thermal Cycler Program Notes  | Multiline Text | Read Only   | <p>Default =<br>Choose the preheated lid option and set to 100°C<br>Reaction volume is 38 µl<br>30°C for 10 minutes<br>Hold at 4°C</p>                                                                                                                                      |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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 3: Clean Up Fragments (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Clean Up Fragments - RNA Prep v1.0
* Step Type = Standard
* Derived Sample Generation = Fixed, 1
* Naming Convention = {InputItemName}
* Reagent Kits
  * Agencourt AMPure XP, 5 ml
    * Supplier = Beckman Coulter
    * Catalog Number = A63880
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-afac16943e1c216039922088bcbe75ca32985830%2Frna-ribo-protocol2-step3-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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** |
  | ------------------ | -------------- | ----------- | ----------------------------------------- |
  | 80% EtOH Prep Date | Date           |             |                                           |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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 4: Amplify Library (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Amplify Library - RNA Prep v1.0
* Step Type = Add Labels
* Derived Sample Generation = Fixed, 1
* Naming Convention = {InputItemName}-{AppliedReagentLabels}
* Reagent Kits
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-7090a0c6eda6a3453fd6b6313acfc90224b992ad%2Frna-ribo-protocol2-step4-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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       |             |                                           |

#### Add Labels

* Label Groups
  * IDT for Illumina RNA UD Indexes Set A Ligation
  * IDT for Illumina RNA UD Indexes Set A-D Ligation
  * IDT for Illumina RNA UD Indexes Set B Ligation
  * IDT for Illumina RNA UD Indexes Set C Ligation
  * IDT for Illumina RNA UD Indexes Set D Ligation
  * Illumina RNA UD Indexes Set A B C D Ligation
  * Illumina RNA UD Indexes Set A Ligation
  * Illumina RNA UD Indexes Set B Ligation
  * Illumina RNA UD Indexes Set C Ligation
  * Illumina RNA UD Indexes Set D Ligation

#### Record Details

* Step Data (Master Step Fields)

  | **Field Name**                       | **Field Type** | **Options**    | **Additional Options and Dropdown Items**                                                                                                                                                                                                                                |
  | ------------------------------------ | -------------- | -------------- | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------ |
  | PCR Cycles                           | Text Dropdown  | Custom Entries | <ul><li><p>Presets</p><ul><li>15</li><li>13</li><li>10</li></ul></li><li>Default = 15</li></ul>                                                                                                                                                                          |
  | Recommendations for PCR Cycle Number | Multiline Text | Read Only      | <ul><li>Default =<br>Use 15 cycles for 10 ng of input DNA<br>Use 13 cycles for 100 ng of input DNA<br>Use 10 cycles for 1000 ng of input DNA</li></ul>                                                                                                                   |
  | Thermal Cycler Program               | Text           |                | <ul><li>Default = PCR</li></ul>                                                                                                                                                                                                                                          |
  | Thermal Cycler Program Notes         | Multiline Text | Read Only      | <ul><li>Default =<br>Choose the preheat lid option and set to 100°C<br>Reaction volume is 50 µl u 98°C for 30 seconds<br>X cycles of:<br>98°C for 10 seconds<br>60°C for 30 seconds<br>72°C for 30 seconds<br>72°C for 5 minutes<br>Hold at 4°C for ≤ 16 hours</li></ul> |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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: Clean Up Library (Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus v1.1)

* Master Step Name = Clean Library v1.0
* Step Type = Standard
* Derived Sample Generation = Fixed, 1
* Naming Convention = {InputItemName}
* Reagent Kits
  * Agencourt AMPure XP, 5 ml
    * Supplier = Beckman Coulter
    * Catalog Number = A63880
  * Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus
    * Supplier = Illumina
    * Website = [www.illumina.com](https://www.illumina.com)

#### Automations

<figure><img src="https://2084401275-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FfjuebS41N49G1Eh55hP7%2Fuploads%2Fgit-blob-852454d61d62ac569dba5e51e5aecdd5df866133%2Frna-ribo-protocol2-step5-automation.png?alt=media" alt=""><figcaption></figcaption></figure>

<details>

<summary>Set Next Step - Advance</summary>

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

{% code overflow="wrap" %}

```markup
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}"
```

{% endcode %}

</details>

#### 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      | 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** |
  | ------------------ | -------------- | ----------- | ----------------------------------------- |
  | 80% EtOH Prep Date | Date           |             |                                           |
* Step File Placeholders
  * Log - Automatically attached
* Sample Table
  * Sample Display Default = Collapse
  * 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       |             |                                           |


---

# Agent Instructions: Querying This Documentation

If you need additional information that is not directly available in this page, you can query the documentation dynamically by asking a question.

Perform an HTTP GET request on the current page URL with the `ask` query parameter:

```
GET https://help.connected.illumina.com/clarity-lims/library-prep/other-rna-protocols/rna-ligation-ribo-v1.1.md?ask=<question>
```

The question should be specific, self-contained, and written in natural language.
The response will contain a direct answer to the question and relevant excerpts and sources from the documentation.

Use this mechanism when the answer is not explicitly present in the current page, you need clarification or additional context, or you want to retrieve related documentation sections.