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Sense-total-RNA

SENSE Total RNA-Seq Library Prep Kit

Company:
  • Strand-specificity >99.9 %
  • Ready-to-sequence libraries in 3.5 hours
  • 0.5 ng of rRNA-depleted or poly(A) selected RNA input
  • incl. from low-quality RNA and FFPE samples

Product Description

SENSE Total RNA-Seq is a strand-specific library prep kit for accurate gene expression profiling, whole transcriptome sequencing, discovery, and quantification of antisense transcripts and overlapping genes.
The typical input amount of poly(A) or rRNA-depleted RNA is 0.5 ng – 500 ng.

Streamlined Ribosomal RNA Depletion

For efficient utilization of RNA-Seq reads highly abundant ribosomal RNA should be eliminated prior library preparation. SENSE Total RNA-Seq Kit is available in combination with RiboCop rRNA Depletion Kit. This is a complete solution for RNA-Seq library preparation, starting with total RNA and finishing with a ready-to-sequence library.

Libraries for Different Sequencing Read Length

For good representation and even coverage of all transcripts in your experiment the library should have a size suitable for the chosen sequencing read length. The size of SENSE libraries can be adjusted by simply modulating the purification steps

Performance

Superior Strand-Specificity

The strand-displacement stop/ligation technology used in SENSE generates fewer antisense artifacts by omitting RT artifacts, and therefore avoids spurious second strand cDNA synthesis, which may result in the detection of false antisense transcription. Thereby an exceptional (99.9%) strand-specificity and reduced experimental noise is reached, enabling the detection and quantification of antisense transcripts with high confidence.

Simple Multiplexing

With the 96 external barcodes included in the kit libraries can be easily multiplexed.

Rapid Turnaround

NGS-ready libraries can be produced from poly(A) selected or rRNA-depleted RNA samples in under 3.5 hours with less than 50% hands-on time, allowing RNA extraction, library preparation and quality control to be performed in one day.

Workflow

Step 1: Library Generation

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The starting material is either ribo-depleted RNA or Poly(A) selected RNA.
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Library generation starts with the random hybridization of the starter/stopper heterodimer mix to the poly(A) selected or rRNA-depleted RNA.
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A single-tube reverse transcription and ligation reaction extends the starter to the next hybridized heterodimer, where the newly synthesized cDNA insert is ligated to the stopper.
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RNA or cDNA fragmentation is not required as the insert size is dictated by the distance between starter/stopper binding sites.
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During second strand synthesis the RNA is hydrolyzed and the library is converted to double-stranded DNA. The double-stranded library is purified using magnetic beads to adjust the library length and get rid of second strand synthesis reaction components.

Step 2: Library Amplification

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The library amplification is performed to add the complete adaptor sequences required for cluster generation and to produce sufficient material for quality control and sequencing. External barcodes can be added during this step in order to be able to multiplex your samples for the sequencing run.
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The finished library is purified from PCR components which can interfere with quantification.

FAQs

1. How is the exceptional strand-specificity achieved?
2. Why do you use ERCC spike-in controls to determine strandedness?
3. How long does it take to generate SENSE libraries?
4. Do I need to purchase additional kits, e.g., for ribosomal RNA depletion, size selection, library amplification, and purification?
5. What are the input RNA requirements for SENSE Total RNA-Seq?
6. Is the kit suitable for preparation of libraries from degraded RNA or FFPE samples?
7. What equipment is needed for performing the SENSE protocol?
8. Which sequencing platforms are suitable for SENSE libraries?
9. What is the orientation of SENSE reads?
10. Why do I see a second peak in high molecular weight regions (between 1,000 - 9,000 bp) on the Bioanalyzer traces?
11. What is a typical library fragment size?
12. Can the insert size be regulated? Is SENSE suitable for longer read lengths e.g. 2x250 bp?
13. How long are the sequences hybridizing to the mRNA? Should those sequences be trimmed?
14. What level of multiplexing can be provided with SENSE? What barcoding (indexing) system do you use?
15. Are adapters of SENSE libraries platform-specific? Which primers should be used for sequencing?
16. What positive control or standards are you recommending to use?
17. What can I do if my libraries are undercycled?
18. Are there any recommendations for FFPE RNA in SENSE Total RNA-Seq?

Ordering Information

Kit Platform Reactions Cat No.
SENSE Total RNA-Seq Library Prep Kit without Ribocop Illumina 8 preps 009.08
24 preps 009.24
96 preps 009.96
SENSE Total RNA-Seq Library Prep Kit without Ribocop Illumina 8 preps 042.08
24 preps 042.24
96 preps 042.96

Downlaods

User Guide Update 24.05.2016 (Recommendations for low input and FFPE RNA; Indication of safe stopping points.)
PCR Add-on Kit for Illumina Instruction Manual
Product Flyer
External Barcodes Overview

Material Safety Datasheets

MSDS information for SENSE Total RNA-Seq Library Prep Kit for Illumina
If you need more information about our products, please contact us through info@moldiag.in