Sensitivity meets Speed
Generate stranded mRNA libraries in under 5 hours with fewer handling and cleanup steps. Designed with automation in mind, the kit comes with generous overages and leverages on-bead washing steps during poly(A) selection to reduce resuspension and wash times, as well as consumable use.
Use a single, flexible workflow to process a broad RNA input range (2.5 ng to 1 µg), without the need for specialized low-input solutions that fail to maintain strand origin information. The Watchmaker mRNA Library Prep Kit delivers highly complex libraries for superior gene detection sensitivity — even with challenging low-input samples.
Key Features and Benefits
- Construct stranded mRNA libraries in under 5 hours - enabling completion in a single workday
- Improve library complexity and gene detection sensitivity with challenging low-input samples (down to 2.5 ng)
- Easily automate with on-magnet washes, fewer cleanup and handling steps, as well as generous overages
- Leverage a novel reverse transcriptase tailored specifically for RNA sequencing
Applications
- Gene expression analysis
- Gene fusion detection
- Isoform/alternative splicing analysis
- Novel transcript discovery
- Single nucleotide variant (SNV) detection
- Pathway analysis
Simple, automatable workflow
Figure 2. A simple, rapid workflow for scalable sample processing. The Watchmaker mRNA Library Prep Kit combines and shortens reaction steps and has fewer cleanups than other commercially available kits. Additionally, it utilizes “on-magnet” bead washing during mRNA capture to omit the need for pipetting to resuspend during washes, as well as reduce magnetting time. Generous reagent overages are included to meet dead volume requirements of liquid handlers.
Key Performance Data
Get more information out of challenging low-input samples
RNA library preparation efficiencies plummet with template-limited samples. This creates a complexity bottleneck and makes it difficult to derive useful biological insight from these libraries. The Watchmaker mRNA Library Prep Kit is specifically optimized for improved performance with low-inputs and delivers superior library complexity in the form of increased unique gene detection.
Figure 3. Increase library complexity and gene ... MORE
Get more information out of challenging low-input samples
Figure 3. Increase library complexity and gene detection sensitivity with low-inputs. RNA extracted from breast tissue (RIN 7) was used to prepare libraries in quadruplicate from a range of RNA mass inputs using the Watchmaker mRNA Library Prep Kit, KAPA mRNA HyperPrep Kit, and the NEBNext® Ultra™ II Directional RNA Library Prep Kit with poly(A) mRNA Enrichment. Supplier recommendations were used for each workflow. KAPA failed to produce libraries at 2.5 ng. Libraries were downsampled to 5M reads. The Watchmaker solution detects significantly more unique genes with low-input samples and an equivalent number of genes when RNA mass is not limiting (with a much-improved workflow). Unique genes were defined as those supported by greater than or equal to 5 unique raw reads.
Maximize sequencing economy
Uninformative ribosomal RNA (rRNA) and duplicate reads effectively limit the capacity of a sequencing run. Reduce the number of reads wasted to rRNA contamination and duplicates with a library prep solution that provides excellent mRNA enrichment and library prep efficiency across a 400-fold input range (2.5 ng to 1 µg). The wide input range enables the use of a single workflow without needing specialized low-input solutions that don't maintain strand origin information.
Maximize sequencing economy
Better preserve gene expression data integrity with low inputs
An overarching goal in transcriptome sequencing is to generate data that is a perfect representation of the biological information contained within a sample.
Many commercially available mRNA library preps struggle to support a wide range of input amounts, resulting in gene expression data distortion. In particular, these solutions often lose relative coverage of lower abundance genes compared to moderate or highly expressed ones – meaning the data output is not representative of the original sample.
The Watchmaker mRNA Library Prep Kit delivers quantitative expression information across a broad input range to provide confidence in results.
Figure 5. Reduce data distortion with ... MORE
Better preserve gene expression data integrity with low inputs
Figure 5. Reduce data distortion with low inputs. RNA extracted from breast tissue (RIN 7) was used to prepare libraries in quadruplicate from a range of RNA mass inputs using the Watchmaker mRNA Library Prep Kit, KAPA mRNA HyperPrep Kit, and the NEBNext® Ultra™ II Directional RNA Library Prep Kit with poly(A) mRNA Enrichment. Supplier recommendations were used for each workflow. Libraries were downsampled to 5M reads. Differential expression analysis between averaged 100 ng (control) and 10 ng (test) libraries was performed using DESeq2 and a raw read cutoff of 5 to call a gene. Results indicate that other vendors lose representation of low abundance genes at 10 ng, while the Watchmaker solution does not. Additionally, Watchmaker libraries exhibit fewer differentially expressed genes between the 10 and 100 ng libraries, indicating better overall agreement in gene expression profiles.
Improve base coverage — even across GC-rich regions
Even transcript coverage uniformity is critical for sensitive applications such as alternative splicing and isoform analysis. The Watchmaker mRNA Library Prep Kit provides consistent coverage uniformity across RNA inputs and leverages Equinox® DNA Polymerase to provide improved coverage of GC-rich regions.
Improve base coverage - even across GC-rich regions
BROCHURES
mRNA Library Prep Kit Brochure
Fusion Detection Gene Expression Analysis Isoform Detection/Alternative Splicing Long Noncoding RNA Analysis mRNA Sequencing Novel Transcript Discovery SNV/SNP/InDel Detection Element Illumina Singular Ultima RNA Blood Dried Blood Spot RNA NGS mRNA Library Prep RNA Library PrepThe Watchmaker mRNA Library Prep Kit creates stranded mRNA-seq libraries in under 5 hours with 2.5 ng–1 µg RNA, offering automation-ready, streamlined workflows and high sensitivity.
TECHNICAL LITERATURE
Maximizing RNA sequencing economy from blood samples using a combined mRNA capture and globin depletion workflow
Gene Expression Analysis Illumina RNA Blood mRNA Library Prep RNA Library Prep with Polaris DepletionThis technical note presents an optimized workflow that combines mRNA capture with globin and rRNA depletion to maximize sequencing efficiency from blood-derived RNA. By reducing non-coding and globin transcript content, this approach improves mRNA enrichment, decreases duplication rates, and enhances gene detection and transcript coverage uniformity across varied RNA inputs.
AUTOMATION RESOURCES
Automated Watchmaker mRNA Library Prep Kit on Sciclone® G3 NGSx
Lab Automation RNA Cell Line RNA NGS mRNA Library Prep RevvityThis application note details the automation of the Watchmaker mRNA Library Prep Kit on the Sciclone® G3 NGSx workstation, enabling the preparation of up to 96 libraries in approximately 5 to 6 hours. The automated workflow reduces hands-on time and minimizes errors, accommodating various sample types and input amounts. The system offers flexibility in bead cleanup ratios and adapter options, enhancing reproducibility and efficiency in mRNA sequencing applications.
POSTERS
Surveying the RNA-seq landscape: A multi-workflow comparison for mRNA and whole transcriptome sequencing applications
Gene Expression Analysis mRNA Sequencing Whole Transcriptome/Total RNA Sequencing Element Illumina Singular RNA Blood Cell Line FFPE Frozen Tissue RNA NGS mRNA Library Prep RNA Library Prep RNA Library Prep with Polaris DepletionThis poster presents a comparative analysis of commercially available stranded RNA library preparation kits for mRNA and whole transcriptome sequencing, focusing on their performance with low-input and degraded samples, such as FFPE. Additionally, it explores compatibility with both Element and Singular short-read sequencing platforms.
PROTOCOLS
Watchmaker mRNA Library Prep Kit User Guide
RNA RNA NGS mRNA Library PrepProtocol for Watchmaker mRNA Library Prep Kit, which enables streamlined preparation of stranded mRNA sequencing libraries from 2.5 ng to 1000 ng of intact total eukaryotic RNA, enriching for polyadenylated mRNAs and converting them into adapter-ligated, double-stranded cDNA fragments.
| RNA input range | 2.5 ng - 1 µg |
|---|---|
| Watchmaker mRNA Capture Kit contents | mRNA Capture Beads Bead Prep Buffer Capture Buffer Final Wash Buffer |
| Watchmaker RNA Library Prep Kit contents | FFPE Treatment Buffer Frag & Prime Buffer 1st Strand Buffer 1st Strand Enzyme 2nd Strand Enzyme 2nd Strand Buffer Ligation Enzyme Ligation Buffer Equinox® Library Amplification Master Mix (2X) P5/P7 Primer Mix (10X) |
| Shipping conditions | Ice packs |
| Storage | mRNA Capture Kit: 4°C RNA Library Prep Kit: -20°C ± 5°C |
| Shelf life | ≥ 6 months |
What applications is the Watchmaker mRNA Library Prep Kit suitable for?
The Watchmaker mRNA Library Prep Kit is ideally suited for:
- Gene expression analysis
- Gene fusion detection
- Isoform/alternative splicing analysis
- Novel transcript discovery
- Single nucleotide variant (SNV) detection
- Pathway analysis
Which species are the Watchmaker mRNA Library Prep Kits compatible with?
The Watchmaker mRNA Library Prep Kit is compatible with eukaryotic species.
Why use mRNA enrichment?
mRNA enrichment focuses sequencing on coding transcripts. If your application only requires coverage of coding transcripts, then mRNA enrichment provides improved sequencing economy by focusing on transcripts you care about.
Does the Watchmaker mRNA Library Prep Kit provide strand orientation information?
Yes, the workflow provides stranded RNA-Seq libraries.
Does this product support long-read sequencing?
We use random primers to prime during first strand synthesis, which does not support full-length cDNA generation. This makes it unlikely to be compatible with-long read applications.
What sample types are compatible with the Watchmaker mRNA Library Prep Kit?
Intact (RIN >7) RNA ranging from 2.5 - 1000 ng is compatible with the Watchmaker mRNA Library Prep Kit. This includes RNA derived from tissues and cell lines, suspended in 50 µl of RNase-free water.
This kit is not compatible with degraded samples, such as FFPE. To prepare libraries from degraded inputs, please use the Watchmaker RNA Library Prep Kit with Polaris® Depletion.
For inputs outside this range, please contact support@watchmakergenomics.com.
What are the RNA purity requirements?
RNA inputs should be free from contaminating DNA. If the total RNA contains DNA, remove the contamination by incubating with DNase I (not supplied with kit). RNA should be suspended in RNase-free water and be free of salts (e.g., Mg2+, or guanidinium salts), chelating agents (e.g., EDTA or EGTA), and organics (e.g., phenol or ethanol).
Can this kit be used to detect non-coding RNAs?
This kit enriches for coding RNA. To detect non-coding RNAs, please use the Watchmaker RNA Library Prep Kit with Polaris Depletion.
Which adapters are the Watchmaker mRNA Library Prep Kit compatible with?
This kit is compatible with adapters that have a 3' overhanging T to facilitate adapter orientation during dscDNA ligation. Note that adapter quality impacts overall library preparation efficiency. Ensure that adapters are adequately duplexed and at the appropriate concentration prior to use. For a list of commercially available adapters and more detailed adapter recommendations, please contact support@watchmakergenomics.com.
Is this kit compatible with other (non-Illumina) sequencing platforms?
This kit has been demonstrated to be compatible with Element, Singular and Ultima sequencing platforms.
Is vortex mixing safe for reactions containing RNA and/or enzyme?
Vortex mixing is recommended for master mix generation and subsequent addition to sample. Pipette mixing is an acceptable alternative so long as care is taken to ensure a completely homogeneous reaction.
What is the shelf life and recommended storage conditions?
The Watchmaker mRNA Library Prep Kit has a shelf life of 6 months upon receipt. The mRNA capture Kit should be stored at 2 - 8°C and the RNA Library Prep Kit at -25°C to -15°C. Both kits should be handled under recommended conditions.
Which modules are included in the Watchmaker mRNA Library Prep Kit?
The mRNA Library Prep Kit (7BK0001) consists of the mRNA capture kit (7K0105) and RNA Library Prep Kit (7K0078). You will receive two boxes, labeled with part numbers 7K0105 and 7K0078.
What is the purpose of the FFPE Treatment Buffer included in the RNA Library Prep module?
The FFPE treatment buffer in the RNA Library Prep Kit box is not used as part of the mRNA Library Prep Kit workflow. This tube may be removed from the kit and discarded.
What other RNA Library Prep Kits does Watchmaker offer?
The Watchmaker RNA Library Prep Kit with Polaris Depletion removes uninformative ribosomal and globin RNAs allowing for whole transcriptome analysis, including long noncoding RNAs. This kit is compatible with both high quality and degraded RNA samples.
The Watchmaker RNA Library Prep Kit is suitable for total RNA and typically used upstream of target enrichment. This kit is compatible with both high quality and degraded RNA samples.
Can this kit be automated?
The Watchmaker mRNA Library Prep Kit is highly automatable. The kit was developed with automation in mind and 96 reaction kits include generous overages to account for dead volumes on liquid handling platforms. Visit our automation page for a list of scripts already developed or to contact a member of our automation team regarding your platform and needs.
Do you offer custom formats?
Yes! Watchmaker offers customer fills, packaging, concentrations, and labeling - including private label - designed to meet your unique needs. We offer flexible terms to serve organizations of any size, and our right-sized processes enable rapid turnaround time on customization. Please contact sales@watchmakergenomics.com to learn more about our capabilities.
Are you ISO 13485-certified?
Yes! Our Quality Management System has achieved ISO 13485:2016 Certification. Our certificate has been awarded for the design, development, manufacture, contract manufacture, and support of high performing reagents for genomics applications in medical research. Download the certificate here.
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| Description | 24 rxn | 96 rxn | 384 rxn | |
|---|---|---|---|---|
| Watchmaker mRNA Library Prep Kit incl. reagents required for mRNA capture, RNA library prep, and amplification | 7BK0001-024 | 7BK0001-096 | 7BK0001-384 | Request a quote |
Please contact sales@watchmakergenomics.com to inquire about custom kit configurations.
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