BioMicroCenter:NanoPore Library Prep - Revision history

Asoberan: Redirect to consolidated BioMicroCenter:LongRead page

2026-04-21T01:49:35Z

Redirect to consolidated BioMicroCenter:LongRead page

← Older revision		Revision as of 01:49, 21 April 2026
Line 1:		Line 1:
	~~{{BioMicroCenter}}~~		#REDIRECT [[BioMicroCenter:LongRead]]
	~~<br><br>~~
	~~Oxford Nanopore Technologies offers a broad variety of methodologies for preparing libraries for sequencing. Pricing includes quality control prior to prep.~~
	~~Large batches will be undertaken on a custom basis; please email biomicro@mit.edu for more information.~~
	~~{\| border=1~~
	~~! Summary~~
	~~! Method~~
	~~! Input~~
	\|-
	~~\| LSK114~~
	~~\| Ligation-based~~
	~~\| 1 ug dsDNA~~
	\|-
	~~\| RBK114~~
	~~\|Tagmentation-based~~
	~~\| 400 ng dsDNA~~
	\|-
	~~\| RNA004~~
	~~\|Ligation-based~~
	~~\| 500 ng polyA+ mRNA (not totalRNA)~~
	\|}
	~~<BR><BR>~~

	~~== dsDNA: SQK-LSK114 ==~~
	{\|
	~~\|- style="vertical-align: top;"~~
	~~\|style="width: 350px;"\|~~
	~~{\| class="wikitable" border=1~~
	~~!Parameter~~
	~~!Requirements~~
	\|-
	~~\|INPUT \|\| Clean double-stranded DNA <BR> 1000 ng+ <BR> >10uL~~
	\|-
	~~\|INCLUDED \|\| Initial QC by UV-vis measurement <BR> Library preparation <BR> Final QC by Qubit~~
	\|-
	~~\|SUBMISSION \|\| MIT - [https://mit.ilabsolutions.com/service_item/new/3381?spt_id=3863&tab=services ilabs] <BR> External - [[BioMicroCenter:Forms\|form]]~~
	\|-
	~~\|UNIT \|\| Per sample~~
	\|-
	~~\|PRICING \|\|[[BioMicroCenter:Pricing#LONG_READ_LIBRARIES\|LINK]]~~
	\|}
	\|
	The BioMicro Center utilizes the SQK-LSK114 kit for most dsDNA library preparation. This kit requires 1 ug of dsDNA and can accommodate a variety of fragment sizes. The enzymes are purchased directly from New England Biolabs or Kapa/Roche. DNA fragments are repaired, A-tailed and ligated to adapters. The ligated products are size-selected using SPRI beads. Prepared libraries can be quantified by Qubit prior to loading. SQK-NBD114 is an extension of this kit which allows for ligation of barcodes prior to pooling and Nanopore adaptor ligation of the pool for sequencing.
	~~<br>~~
	\|}

	~~== dsDNA: SQK-RBK114 ==~~
	{\|
	~~\|- style="vertical-align: top;"~~
	~~\|style="width: 350px;"\|~~
	~~{\| class="wikitable" border=1~~
	~~!Parameter~~
	~~!Requirements~~
	\|-
	~~\|INPUT \|\| Clean double-stranded DNA <BR> 400 ng+ <BR> >30 kbp <br> >10uL~~
	\|-
	~~\|INCLUDED \|\| Initial QC by UV-vis and FemtoPulse <BR> Library preparation <BR> Final QC by Qubit~~
	\|-
	~~\|SUBMISSION \|\| MIT - [https://mit.ilabsolutions.com/service_item/new/3381?spt_id=3863&tab=services ilabs] <BR> External - [[BioMicroCenter:Forms\|form]]~~
	\|-
	~~\|UNIT \|\| Per sample~~
	\|-
	~~\|PRICING \|\|[[BioMicroCenter:Pricing~~#~~LONG_READ_LIBRARIES\|LINK]]~~
	\|}
	\|
	The BioMicro Center utilizes the SQK-RBK114 kit for tagmentation-based library preparation for dsDNA of large size distributions. Oxford recommends sizes greater than 30 kbp with input of at least 400 ng. The enzymes are bought directly from New England Biolabs. Input is tagmented with adapters and the products are size selected using SPRI beads. Prepared library will be quantified by Qubit prior to loading.
	~~<br><br>~~
	This kit can also be used when users are interested in obtaining the longest reads possible. For this protocol, micrograms of clean genomic DNA must be used. No size selection is performed. The PromethION P2S has sequenced reads with lengths of 300 kbp+: however, these long reads are rare outliers and the overall read amount for the flowcell in use will be lowered. Higher Molecular Weight DNA seems to kill the nanopores faster and requires a large amount of computing power from the sequencer, reducing the real time speed of basecalling and delaying data delivery.
	\|}

	~~== RNA: SQK-RNA004 ==~~
	{\|
	~~\|- style="vertical-align: top;"~~
	~~\|style="width: 350px;"\|~~
	~~{\| class="wikitable" border=1~~
	~~!Parameter~~
	~~!Requirements~~
	\|-
	~~\|INPUT \|\| Clean mRNA <BR> 500 ng+ <br> >10uL~~
	\|-
	~~\|INCLUDED \|\| Initial QC by UV-vis and Fragment Analyzer <BR> Library preparation <BR> Final QC by Qubit~~
	\|-
	~~\|SUBMISSION \|\| MIT - [https://mit.ilabsolutions.com/service_item/new/3381?spt_id=3863&tab=services ilabs] <BR> External -~~ [[BioMicroCenter:~~Forms\|form]]~~
	\|-
	~~\|UNIT \|\| Per sample~~
	\|-
	~~\|PRICING \|\|[[BioMicroCenter:Pricing#LONG_READ_LIBRARIES\|LINK~~]]
	\|}
	\|
	The BioMicro Center utilizes the SQK-RNA004 kit for library preparation with RNA. The enzymes are purchased directly from New England Biolabs. The typical workflow uses first strand synthesis to create a site for ligation--this step can be omitted if necessary, but significantly reduces read amount. Input is ligated with adapters and the products are size selected using SPRI beads. Prepared library can be quantified by Qubit prior to loading.
	~~<br><br>~~
	~~SPRI bead cleans, poly-T bead selection, gel separation and other methods can be added to further select the population of RNA prior to library preparation and sequencing.~~
	\|}

Hilo1: /* dsDNA: SQK-RBK114 */

2025-05-20T19:25:37Z

dsDNA: SQK-RBK114

← Older revision		Revision as of 19:25, 20 May 2025
Line 66:		Line 66:
	The BioMicro Center utilizes the SQK-RBK114 kit for tagmentation-based library preparation for dsDNA of large size distributions. Oxford recommends sizes greater than 30 kbp with input of at least 400 ng. The enzymes are bought directly from New England Biolabs. Input is tagmented with adapters and the products are size selected using SPRI beads. Prepared library will be quantified by Qubit prior to loading.		The BioMicro Center utilizes the SQK-RBK114 kit for tagmentation-based library preparation for dsDNA of large size distributions. Oxford recommends sizes greater than 30 kbp with input of at least 400 ng. The enzymes are bought directly from New England Biolabs. Input is tagmented with adapters and the products are size selected using SPRI beads. Prepared library will be quantified by Qubit prior to loading.
	<br><br>		<br><br>
	This kit can also be used when users are interested in obtaining the longest reads possible. For this protocol, micrograms of clean genomic DNA must be used. No size selection is performed. The PromethION ~~P24~~ has sequenced reads with lengths of 300 kbp+: however, these long reads are rare outliers and the overall read amount for the flowcell in use will be lowered. ~~Longer~~ DNA ~~and/or carryover~~ seems to kill the nanopores faster and requires a large amount of computing power from the sequencer, reducing the ~~amount~~ of ~~flowcells that can be simultaneously loaded on the sequencer~~.		This kit can also be used when users are interested in obtaining the longest reads possible. For this protocol, micrograms of clean genomic DNA must be used. No size selection is performed. The PromethION P2S has sequenced reads with lengths of 300 kbp+: however, these long reads are rare outliers and the overall read amount for the flowcell in use will be lowered. Higher Molecular Weight DNA seems to kill the nanopores faster and requires a large amount of computing power from the sequencer, reducing the real time speed of basecalling and delaying data delivery.
	\|}		\|}

Hilo1: /* dsDNA: SQK-LSK114 */

2025-05-20T19:20:25Z

dsDNA: SQK-LSK114

← Older revision		Revision as of 19:20, 20 May 2025
Line 41:		Line 41:
	\|}		\|}
	\|		\|
	The BioMicro Center utilizes the SQK-LSK114 kit for most dsDNA library preparation. This kit requires 1 ug of dsDNA and can accommodate a variety of fragment sizes. The enzymes are purchased directly from New England Biolabs or Kapa/Roche. DNA fragments are repaired, A-tailed and ligated to adapters. The ligated products are size-selected using SPRI beads. Prepared libraries can be quantified by Qubit prior to loading. SQK-NBD114 is an extension of this kit which allows for ligation of barcodes prior to pooling and Nanopore adaptor ligation for sequencing.		The BioMicro Center utilizes the SQK-LSK114 kit for most dsDNA library preparation. This kit requires 1 ug of dsDNA and can accommodate a variety of fragment sizes. The enzymes are purchased directly from New England Biolabs or Kapa/Roche. DNA fragments are repaired, A-tailed and ligated to adapters. The ligated products are size-selected using SPRI beads. Prepared libraries can be quantified by Qubit prior to loading. SQK-NBD114 is an extension of this kit which allows for ligation of barcodes prior to pooling and Nanopore adaptor ligation of the pool for sequencing.
	<br>		<br>
	\|}		\|}

Hilo1: /* dsDNA: SQK-LSK114 */

2025-05-20T19:20:04Z

dsDNA: SQK-LSK114

← Older revision		Revision as of 19:20, 20 May 2025
Line 41:		Line 41:
	\|}		\|}
	\|		\|
	The BioMicro Center utilizes the SQK-LSK114 kit for most dsDNA library preparation. This kit requires 1 ug of dsDNA and can accommodate a variety of fragment sizes. The enzymes are purchased directly from New England Biolabs or Kapa/Roche. DNA fragments are repaired, A-tailed and ligated to adapters. The ligated products are size-selected using SPRI beads. Prepared libraries can be quantified by Qubit prior to loading.		The BioMicro Center utilizes the SQK-LSK114 kit for most dsDNA library preparation. This kit requires 1 ug of dsDNA and can accommodate a variety of fragment sizes. The enzymes are purchased directly from New England Biolabs or Kapa/Roche. DNA fragments are repaired, A-tailed and ligated to adapters. The ligated products are size-selected using SPRI beads. Prepared libraries can be quantified by Qubit prior to loading. SQK-NBD114 is an extension of this kit which allows for ligation of barcodes prior to pooling and Nanopore adaptor ligation for sequencing.
	<br>		<br>
	\|}		\|}

Hilo1 at 19:18, 20 May 2025

2025-05-20T19:18:26Z

← Older revision		Revision as of 19:18, 20 May 2025
Line 12:		Line 12:
	\| 1 ug dsDNA		\| 1 ug dsDNA
	\|-		\|-
	\| ~~RAD004~~		\| RBK114
	\|Tagmentation-based		\|Tagmentation-based
	\| 400 ng dsDNA		\| 400 ng dsDNA
	\|-		\|-
	\| ~~RNA002~~		\| RNA004
	\|Ligation-based		\|Ligation-based
	\| 500 ng polyA+ mRNA (not totalRNA)		\| 500 ng polyA+ mRNA (not totalRNA)

Hilo1: /* RNA: SQK-RNA004 */

2025-05-20T19:16:49Z

RNA: SQK-RNA004

← Older revision		Revision as of 19:16, 20 May 2025
Line 88:		Line 88:
	\|}		\|}
	\|		\|
	The BioMicro Center utilizes the SQK-~~RNA002~~ kit for library preparation with RNA. The enzymes are purchased directly from New England Biolabs. The typical workflow uses first strand synthesis to create a site for ligation--this step can be omitted if necessary, but significantly reduces read amount. Input is ligated with adapters and the products are size selected using SPRI beads. Prepared library can be quantified by Qubit prior to loading.		The BioMicro Center utilizes the SQK-RNA004 kit for library preparation with RNA. The enzymes are purchased directly from New England Biolabs. The typical workflow uses first strand synthesis to create a site for ligation--this step can be omitted if necessary, but significantly reduces read amount. Input is ligated with adapters and the products are size selected using SPRI beads. Prepared library can be quantified by Qubit prior to loading.
	<br><br>		<br><br>
	SPRI bead cleans, poly-T bead selection, gel separation and other methods can be added to further select the population of RNA prior to library preparation and sequencing.		SPRI bead cleans, poly-T bead selection, gel separation and other methods can be added to further select the population of RNA prior to library preparation and sequencing.
	\|}		\|}

Hilo1 at 19:16, 20 May 2025

2025-05-20T19:16:30Z

← Older revision		Revision as of 19:16, 20 May 2025
Line 45:		Line 45:
	\|}		\|}

	== dsDNA: SQK-~~RAD004~~ ==		== dsDNA: SQK-RBK114 ==
	{\|		{\|
	\|- style="vertical-align: top;"		\|- style="vertical-align: top;"
Line 64:		Line 64:
	\|}		\|}
	\|		\|
	The BioMicro Center utilizes the SQK-~~RAD004~~ kit for tagmentation-based library preparation for dsDNA of large size distributions. Oxford recommends sizes greater than 30 kbp with input of at least 400 ng. The enzymes are bought directly from New England Biolabs. Input is tagmented with adapters and the products are size selected using SPRI beads. Prepared library will be quantified by Qubit prior to loading.		The BioMicro Center utilizes the SQK-RBK114 kit for tagmentation-based library preparation for dsDNA of large size distributions. Oxford recommends sizes greater than 30 kbp with input of at least 400 ng. The enzymes are bought directly from New England Biolabs. Input is tagmented with adapters and the products are size selected using SPRI beads. Prepared library will be quantified by Qubit prior to loading.
	<br><br>		<br><br>
	This kit can also be used when users are interested in obtaining the longest reads possible. For this protocol, micrograms of clean genomic DNA must be used. No size selection is performed. The PromethION P24 has sequenced reads with lengths of 300 kbp+: however, these long reads are rare outliers and the overall read amount for the flowcell in use will be lowered. Longer DNA and/or carryover seems to kill the nanopores faster and requires a large amount of computing power from the sequencer, reducing the amount of flowcells that can be simultaneously loaded on the sequencer.		This kit can also be used when users are interested in obtaining the longest reads possible. For this protocol, micrograms of clean genomic DNA must be used. No size selection is performed. The PromethION P24 has sequenced reads with lengths of 300 kbp+: however, these long reads are rare outliers and the overall read amount for the flowcell in use will be lowered. Longer DNA and/or carryover seems to kill the nanopores faster and requires a large amount of computing power from the sequencer, reducing the amount of flowcells that can be simultaneously loaded on the sequencer.

Hilo1 at 19:14, 20 May 2025

2025-05-20T19:14:11Z

← Older revision		Revision as of 19:14, 20 May 2025
Line 69:		Line 69:
	\|}		\|}

	== RNA: SQK-~~RNA002~~ ==		== RNA: SQK-RNA004 ==
	{\|		{\|
	\|- style="vertical-align: top;"		\|- style="vertical-align: top;"

imported>Stuart S. Levine at 00:53, 5 October 2023

2023-10-05T00:53:20Z

New page

{{BioMicroCenter}}
 
Oxford Nanopore Technologies offers a broad variety of methodologies for preparing libraries for sequencing. Pricing includes quality control prior to prep.
Large batches will be undertaken on a custom basis; please email biomicro@mit.edu for more information.
{| border=1
! Summary
! Method
! Input
|-
| LSK114
| Ligation-based
| 1 ug dsDNA
|-
| RAD004
|Tagmentation-based
| 400 ng dsDNA
|-
| RNA002
|Ligation-based
| 500 ng polyA+ mRNA (not totalRNA)
|}
 

== dsDNA: SQK-LSK114 ==
{|
|- style="vertical-align: top;"
|style="width: 350px;"|
{| class="wikitable" border=1
!Parameter
!Requirements
|-
|INPUT || Clean double-stranded DNA 1000 ng+ >10uL
|-
|INCLUDED || Initial QC by UV-vis measurement Library preparation Final QC by Qubit
|-
|SUBMISSION || MIT - [https://mit.ilabsolutions.com/service_item/new/3381?spt_id=3863&tab=services ilabs] External - [[BioMicroCenter:Forms|form]]
|-
|UNIT || Per sample
|-
|PRICING ||[[BioMicroCenter:Pricing#LONG_READ_LIBRARIES|LINK]]
|}
|
The BioMicro Center utilizes the SQK-LSK114 kit for most dsDNA library preparation. This kit requires 1 ug of dsDNA and can accommodate a variety of fragment sizes. The enzymes are purchased directly from New England Biolabs or Kapa/Roche. DNA fragments are repaired, A-tailed and ligated to adapters. The ligated products are size-selected using SPRI beads. Prepared libraries can be quantified by Qubit prior to loading.
 
|}

== dsDNA: SQK-RAD004 ==
{|
|- style="vertical-align: top;"
|style="width: 350px;"|
{| class="wikitable" border=1
!Parameter
!Requirements
|-
|INPUT || Clean double-stranded DNA 400 ng+ >30 kbp >10uL
|-
|INCLUDED || Initial QC by UV-vis and FemtoPulse Library preparation Final QC by Qubit
|-
|SUBMISSION || MIT - [https://mit.ilabsolutions.com/service_item/new/3381?spt_id=3863&tab=services ilabs] External - [[BioMicroCenter:Forms|form]]
|-
|UNIT || Per sample
|-
|PRICING ||[[BioMicroCenter:Pricing#LONG_READ_LIBRARIES|LINK]]
|}
|
The BioMicro Center utilizes the SQK-RAD004 kit for tagmentation-based library preparation for dsDNA of large size distributions. Oxford recommends sizes greater than 30 kbp with input of at least 400 ng. The enzymes are bought directly from New England Biolabs. Input is tagmented with adapters and the products are size selected using SPRI beads. Prepared library will be quantified by Qubit prior to loading.
 
This kit can also be used when users are interested in obtaining the longest reads possible. For this protocol, micrograms of clean genomic DNA must be used. No size selection is performed. The PromethION P24 has sequenced reads with lengths of 300 kbp+: however, these long reads are rare outliers and the overall read amount for the flowcell in use will be lowered. Longer DNA and/or carryover seems to kill the nanopores faster and requires a large amount of computing power from the sequencer, reducing the amount of flowcells that can be simultaneously loaded on the sequencer.
|}

== RNA: SQK-RNA002 ==
{|
|- style="vertical-align: top;"
|style="width: 350px;"|
{| class="wikitable" border=1
!Parameter
!Requirements
|-
|INPUT || Clean mRNA 500 ng+ >10uL
|-
|INCLUDED || Initial QC by UV-vis and Fragment Analyzer Library preparation Final QC by Qubit
|-
|SUBMISSION || MIT - [https://mit.ilabsolutions.com/service_item/new/3381?spt_id=3863&tab=services ilabs] External - [[BioMicroCenter:Forms|form]]
|-
|UNIT || Per sample
|-
|PRICING ||[[BioMicroCenter:Pricing#LONG_READ_LIBRARIES|LINK]]
|}
|
The BioMicro Center utilizes the SQK-RNA002 kit for library preparation with RNA. The enzymes are purchased directly from New England Biolabs. The typical workflow uses first strand synthesis to create a site for ligation--this step can be omitted if necessary, but significantly reduces read amount. Input is ligated with adapters and the products are size selected using SPRI beads. Prepared library can be quantified by Qubit prior to loading.
 
SPRI bead cleans, poly-T bead selection, gel separation and other methods can be added to further select the population of RNA prior to library preparation and sequencing.
|}