Ligation sequencing V14 - PCR barcoding (SQK-LSK114 with EXP-PBC001 or EXP-PBC096) (PBC_9182_v114_revM_30Jan2025)

DateVersionLanguageDeviceChanges
5/14/2025PBC_9182_v114_revM_30Jan2025EnglishGridION
May 2025 - In 'Overview', library preparation time was increased to ~155 min to align with Table in 'Steps in the sequencing workflow' where priming and loading of flow cell was increased from 5 min to 10 min. - In sections 3 and 6 'End-prep', step 5, the following was added to all device arms: 'Then cool down to between 4°C and 20°C on the thermal cycler or place the samples on ice.'. - In section 8, 'Priming and loading the MinION and GridION Flow Cell' an important note was added to the bottom of this section stating that when a flow cell is inserted into the MinION Mk1D and the lid is closed, there will be a small gap around the sides of the device. - For the PromethION branch, the step for removing the PromethION Flow Cell out of the fridge was moved to the beginning of the section on 'Priming and loading the PromethION Flow Cell' to enable the flow cell to reach room temperature while the flow cell priming mix is prepared. And an optional action of performing a flow cell check was included.
5/14/2025PBC_9182_v114_revM_30Jan2025EnglishMinION
May 2025 - In 'Overview', library preparation time was increased to ~155 min to align with Table in 'Steps in the sequencing workflow' where priming and loading of flow cell was increased from 5 min to 10 min. - In sections 3 and 6 'End-prep', step 5, the following was added to all device arms: 'Then cool down to between 4°C and 20°C on the thermal cycler or place the samples on ice.'. - In section 8, 'Priming and loading the MinION and GridION Flow Cell' an important note was added to the bottom of this section stating that when a flow cell is inserted into the MinION Mk1D and the lid is closed, there will be a small gap around the sides of the device. - For the PromethION branch, the step for removing the PromethION Flow Cell out of the fridge was moved to the beginning of the section on 'Priming and loading the PromethION Flow Cell' to enable the flow cell to reach room temperature while the flow cell priming mix is prepared. And an optional action of performing a flow cell check was included.
5/14/2025PBC_9182_v114_revM_30Jan2025EnglishPromethION
May 2025 - In 'Overview', library preparation time was increased to ~155 min to align with Table in 'Steps in the sequencing workflow' where priming and loading of flow cell was increased from 5 min to 10 min. - In sections 3 and 6 'End-prep', step 5, the following was added to all device arms: 'Then cool down to between 4°C and 20°C on the thermal cycler or place the samples on ice.'. - In section 8, 'Priming and loading the MinION and GridION Flow Cell' an important note was added to the bottom of this section stating that when a flow cell is inserted into the MinION Mk1D and the lid is closed, there will be a small gap around the sides of the device. - For the PromethION branch, the step for removing the PromethION Flow Cell out of the fridge was moved to the beginning of the section on 'Priming and loading the PromethION Flow Cell' to enable the flow cell to reach room temperature while the flow cell priming mix is prepared. And an optional action of performing a flow cell check was included.
1/30/2025PBC_9182_v114_revM_30Jan2025EnglishGridION
30th January 2025 In "Adapter ligation and clean-up". the instructions for preparing the LFB or SFB have been changed to: "Thaw either Long Fragment Buffer (LFB) or Short Fragment Buffer (SFB) at room temperature and mix by vortexing. Then spin down and keep at room temperature."
1/30/2025PBC_9182_v114_revM_30Jan2025EnglishMinION
30th January 2025 In "Adapter ligation and clean-up". the instructions for preparing the LFB or SFB have been changed to: "Thaw either Long Fragment Buffer (LFB) or Short Fragment Buffer (SFB) at room temperature and mix by vortexing. Then spin down and keep at room temperature."
1/30/2025PBC_9182_v114_revM_30Jan2025EnglishPromethION
30th January 2025 In "Adapter ligation and clean-up". the instructions for preparing the LFB or SFB have been changed to: "Thaw either Long Fragment Buffer (LFB) or Short Fragment Buffer (SFB) at room temperature and mix by vortexing. Then spin down and keep at room temperature."
12/12/2024PBC_9182_v114_revL_12Dec2024EnglishGridION
12 December 2024 - Addition of link to MinION Mk1D IT requirements in "Compatibility of this protocol". - Arrow which was previously at the bottom edge of the flow cell in Image 1b "Priming and loading MinION/GridION Flow Cell" was moved to the middle of the flow cell. - Updated link for flow cell returns in "Flow cell reuse and returns" and removed note for flushing flow cells. - Updated link for MinION temperature control in "Issues during a sequencing run", "Failed to reach target temperature".
12/12/2024PBC_9182_v114_revL_12Dec2024EnglishMinION
12 December 2024 - Addition of link to MinION Mk1D IT requirements in "Compatibility of this protocol". - Arrow which was previously at the bottom edge of the flow cell in Image 1b "Priming and loading MinION/GridION Flow Cell" was moved to the middle of the flow cell. - Updated link for flow cell returns in "Flow cell reuse and returns" and removed note for flushing flow cells. - Updated link for MinION temperature control in "Issues during a sequencing run", "Failed to reach target temperature".
12/12/2024PBC_9182_v114_revL_12Dec2024EnglishPromethION
12 December 2024 - Addition of link to MinION Mk1D IT requirements in "Compatibility of this protocol". - Arrow which was previously at the bottom edge of the flow cell in Image 1b "Priming and loading MinION/GridION Flow Cell" was moved to the middle of the flow cell. - Updated link for flow cell returns in "Flow cell reuse and returns" and removed note for flushing flow cells. - Updated link for MinION temperature control in "Issues during a sequencing run", "Failed to reach target temperature".
8/21/2024PBC_9182_v114_revK_07Mar2023EnglishGridION
21st August 2024 - Updated protocol introduction to new format. - Updated sample input recommendations to 100 ng for gDNA or amplicons. - NEB reagent recommendations for purchase and use updated. - Updated the End-prep method in-line with new recommended reagents. - Updated the Adapter ligation step with new recommended method. - Updated library loading on flow cell recommendations with table for different DNA size.
8/21/2024PBC_9182_v114_revK_07Mar2023EnglishMinION
21st August 2024 - Updated protocol introduction to new format. - Updated sample input recommendations to 100 ng for gDNA or amplicons. - NEB reagent recommendations for purchase and use updated. - Updated the End-prep method in-line with new recommended reagents. - Updated the Adapter ligation step with new recommended method. - Updated library loading on flow cell recommendations with table for different DNA size.
8/21/2024PBC_9182_v114_revK_07Mar2023EnglishPromethION
21st August 2024 - Updated protocol introduction to new format. - Updated sample input recommendations to 100 ng for gDNA or amplicons. - NEB reagent recommendations for purchase and use updated. - Updated the End-prep method in-line with new recommended reagents. - Updated the Adapter ligation step with new recommended method. - Updated library loading on flow cell recommendations with table for different DNA size.
August 21st, 2024PBC_9182_v114_revJ_07Mar2023EnglishPromethion Download 31 July 2024 - Changed reagent order addition in Library preparation/Adapter ligation and clean-up; Ligation Adapter (LA) added before Ligation Buffer (LB) and NEBNext Quick T4 Ligase 07th March 2024 - Updated library loading recommendations to 35-50 fmols.
August 21st, 2024PBC_9182_v114_revJ_07Mar2023EnglishGridion Download 31 July 2024 - Changed reagent order addition in Library preparation/Adapter ligation and clean-up; Ligation Adapter (LA) added before Ligation Buffer (LB) and NEBNext Quick T4 Ligase 07th March 2024 - Updated library loading recommendations to 35-50 fmols.
August 21st, 2024PBC_9182_v114_revJ_07Mar2023EnglishMinion Download 31 July 2024 - Changed reagent order addition in Library preparation/Adapter ligation and clean-up; Ligation Adapter (LA) added before Ligation Buffer (LB) and NEBNext Quick T4 Ligase 07th March 2024 - Updated library loading recommendations to 35-50 fmols.
July 31st, 2024PBC_9182_v114_revI_07Mar2023EnglishPromethion Download 07th March 2024 - Updated library loading recommendations to 35-50 fmols.
July 31st, 2024PBC_9182_v114_revI_07Mar2023EnglishGridion Download 07th March 2024 - Updated library loading recommendations to 35-50 fmols.
July 31st, 2024PBC_9182_v114_revI_07Mar2023EnglishMinion Download 07th March 2024 - Updated library loading recommendations to 35-50 fmols.
September 19th, 2023PBC_9182_v114_revG_07Mar2023EnglishPromethion Download 1st August 2023 - Kit contents layout has been updated - In priming and loading the flow cell, the flow cell priming mix instructions have been updated - Added amplicon tailed primer sequences to the introduction
September 19th, 2023PBC_9182_v114_revG_07Mar2023EnglishGridion Download 1st August 2023 - Kit contents layout has been updated - In priming and loading the flow cell, the flow cell priming mix instructions have been updated - Added amplicon tailed primer sequences to the introduction
September 19th, 2023PBC_9182_v114_revG_07Mar2023EnglishMinion Download 1st August 2023 - Kit contents layout has been updated - In priming and loading the flow cell, the flow cell priming mix instructions have been updated - Added amplicon tailed primer sequences to the introduction
May 3rd, 2023PBC_9182_v114_revF_07Mar2023EnglishPromethion Download May 2023 - Added flow cell priming mix volumes to the "Priming and loading" steps - This kit has moved to released phase.
May 3rd, 2023PBC_9182_v114_revF_07Mar2023EnglishGridion Download May 2023 - Added flow cell priming mix volumes to the "Priming and loading" steps - This kit has moved to released phase.
May 3rd, 2023PBC_9182_v114_revF_07Mar2023EnglishMinion Download May 2023 - Added flow cell priming mix volumes to the "Priming and loading" steps - This kit has moved to released phase.
April 28th, 2023PBC_9182_v114_revE_07Mar2023EnglishPromethion Download April 2023 - Added flow cell priming mix volumes to the "Priming and loading" steps
April 28th, 2023PBC_9182_v114_revE_07Mar2023EnglishGridion Download April 2023 - Added flow cell priming mix volumes to the "Priming and loading" steps
April 28th, 2023PBC_9182_v114_revE_07Mar2023EnglishMinion Download April 2023 - Added flow cell priming mix volumes to the "Priming and loading" steps
March 24th, 2023PBC_9182_v114_revD_07Mar2023EnglishPromethion Download March 2023: - Updated how much DNA is expected and taken forwards from the first end-prep step
March 24th, 2023PBC_9182_v114_revD_07Mar2023EnglishGridion Download March 2023: - Updated how much DNA is expected and taken forwards from the first end-prep step
March 24th, 2023PBC_9182_v114_revD_07Mar2023EnglishMinion Download March 2023: - Updated how much DNA is expected and taken forwards from the first end-prep step
March 17th, 2023PBC_9182_v114_revC_07Mar2023EnglishPromethion Download March 2023: - Added instructions for preparing ONT reagents
March 17th, 2023PBC_9182_v114_revC_07Mar2023EnglishGridion Download March 2023: - Added instructions for preparing ONT reagents
March 17th, 2023PBC_9182_v114_revC_07Mar2023EnglishMinion Download March 2023: - Added instructions for preparing ONT reagents
March 16th, 2023PBC_9182_v114_revB_07Mar2023EnglishPromethion Download March 2023: - Updated input recommendations for gDNA in the "Equipment and consumables" section, as well as in the "End-prep" step.
March 16th, 2023PBC_9182_v114_revB_07Mar2023EnglishGridion Download March 2023: - Updated input recommendations for gDNA in the "Equipment and consumables" section, as well as in the "End-prep" step.
March 16th, 2023PBC_9182_v114_revB_07Mar2023EnglishMinion Download March 2023: - Updated input recommendations for gDNA in the "Equipment and consumables" section, as well as in the "End-prep" step.
March 8th, 2023PBC_9182_v114_revA_07Mar2023EnglishPromethion Download -
March 8th, 2023PBC_9182_v114_revA_07Mar2023EnglishGridion Download -
March 8th, 2023PBC_9182_v114_revA_07Mar2023EnglishMinion Download -