RNA for Illumina®
For use with the Illumina® sequencing platform, RNA libraries are prepared by depletion of abundant RNAs from total RNA, and synthesis of cDNA followed by DNA library preparation steps: repair of 3′ and 5′ ends followed by the addition of a non-templated dA-tail before ligation with an adaptor. Libraries are size selected after adaptor ligation, and amplified via polymerase chain reaction (PCR). For more information, choose either the Ultra™ II Directional or Non-directional RNA Library Preparation Workflow for Illumina tab below.
Abundant RNAs can be removed from total RNA directly using an RNA depletion product specific for the sample species being used, or by enrichment of mRNA by a poly(A)-based method.
For single cell and ultra-low input RNA samples, libraries are constructed with a template-switching method to generate full-length cDNAs, followed by Ultra II library preparation. For more information, choose the Single Cell/Low Input RNA Library Preparation Workflow for Illumina tab below.
Small RNA libraries are constructed using a different workflow, in which adaptors are ligated directly to the small RNA molecules, followed by reverse transcription, PCR amplification and size selection. In the novel NEBNext® workflow, the introduction of the RT primer ahead of ligation of the 5’ adaptor prevents formation of contaminating adaptor-dimers. For more information, choose the Small RNA Library Preparation Workflow for Illumina tab below.
NEBNext reagents are available for each step in RNA and small RNA library preparation workflows for the Illumina platform. Reagents from NEB are available in multiple formats, and a wide range of index primers for multiplexing is available. Each of these reagents has been functionally validated by library preparation followed by Illumina sequencing.
Illumina® is a registered trademark of Illumina, Inc.
Choose Type:
- NEBNext® Magnesium RNA Fragmentation Module Protocol
- Protocol for NEBNext® Ultra™ II Non-Directional RNA Second Strand Synthesis Module (E6111)
- Validate the Library - NEBNext Multiplex Small RNA Sample Prep Set for Illumina (For use with NEB #E7300, #E7330, and #E7580)
- Isolate mRNA using the NEBNext® Poly(A) mRNA Magnetic Isolation Module (E7490)
- NEBNext Multiplex Small RNA Sample Prep Set for Illumina - Library Preparation (E7330)
- Protocol for NEBNext® Ultra™ II Directional RNA Second Strand Synthesis Module (E7550)
- Protocol for Use with Previously Fragmented RNA - NEBNext mRNA First Strand Synthesis Module (E7525)
- Protocol for Use with Total RNA - NEBNext mRNA First Strand Synthesis Module (E7525)
- Library Preparation - NEBNext Multiplex Small RNA Sample Prep Set for Illumina (E7580)
- QC Check and Size Selection using 6% PolyAcrylamide Gel - NEBNext Multiplex Small RNA Sample Prep Set for Illumina (E7300)
- QC Check and Size Selection using 6% PolyAcrylamide Gel (E7330)
- QC Check and Size Selection using AMPure XP Beads - NEBNext Multiplex Small RNA Sample Prep Set for Illumina (E7300)
- QC Check and Size Selection using AMPure XP Beads (E7330)
- QC Check and Size Selection Using Pippin Prep - NEBNext Multiplex Small RNA Sample Prep Set for Illumina (E7300)
- Protocol for use with NEBNext mRNA Library Prep Master Mix Set for Illumina (E6110)
- Protocol for use with NEBNext Poly(A) mRNA Magnetic Isolation Module, NEB #E7490 and NEBNext Ultra RNA Library Prep Kit for Illumina (NEB #E7530)
- Protocol for use with NEBNext rRNA Depletion Kit (Human/Mouse/Rat) (E6310) and NEBNext Ultra Directional RNA Library Prep Kit for Illumina (E7420)
- Protocol for use with NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490) and NEBNext Ultra Directional RNA Library Prep Kit for Illumina (E7420)
- Protocol for use with NEBNext rRNA Depletion Kit (Human/Mouse/Rat), NEB #E6310 and NEBNext Ultra RNA Library Prep Kit for Illumina (NEB #E7530)
- Please see manual (NEB #E6100) for protocols
- Refer to the manual (NEB #E7335) for a list of NEBNext library prep kits that contain protocols for use of NEBNext adaptors and primers.
- Please see manual (NEB #E7535) for protocols
- Protocol for use with Purified mRNA or Ribosome Depleted RNA - NEBNext Ultra Directional RNA Library Prep Kit for Illumina (E7420)
- Protocol for use with Purified mRNA or Ribosome Depleted RNA - NEBNext Ultra RNA Library Prep Kit for Illumina (E7530)
- QC Check and Size Selection Using Pippin Prep (E7330)
- Protocol for use with NEBNext rRNA Depletion Kit (Human/Mouse/Rat) (NEB #E6310)
- Protocol for use with NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490) and NEBNext Ultra II RNA Library Prep Kit for Illumina (E7770, E7775)
- Protocol for use with FFPE RNA, NEBNext rRNA Depletion Kit (Human/Mouse/Rat) (E6310) and NEBNext Ultra II RNA Library Prep Kit for Illumina (E7770, E7775)
- Protocol for use with Purified mRNA or rRNA Depleted RNA and NEBNext Ultra II RNA Library Prep Kit for Illumina (E7770, E7775)
- Protocol for use with rRNA Depleted FFPE RNA and NEBNext Ultra II RNA Library Prep Kit for Illumina (E7770, E7775)
- Protocol for Directional RNA-seq Workflows and NEBNext Ultra II RNA First Strand Synthesis Module (E7771)
- Protocol for Non-directional RNA-seq Workflows and NEBNext Ultra II RNA First Strand Synthesis Module (E7771)
- Appendix A
- Appendix A for use with NEBNext Ultra II RNA First Strand Synthesis Module (E7771)
While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.
For more information about commercial rights, please contact NEB's Global Business Development team at [email protected].
This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.