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  • ‹ Back to RNA-seq
  • Stranded libraries from picogram-input total RNA (v3)
  • Automation-friendly, all-in-one cDNA synthesis and library prep
  • All-in-one cDNA synthesis and library prep from ultra-low RNA inputs
  • Stranded libraries from picogram-input total RNA (v2)
  • Stranded libraries from FFPE inputs (v2)
  • Stranded libraries from 100 ng - 1 ug total RNA
  • Stranded libraries from 100 pg-100 ng total RNA
  • Stranded libraries from picogram-input total RNA (v1)
  • Stranded RNA-seq competitor kit comparison
  • Nonstranded libraries from FFPE inputs
  • Sensitive capture of full-length transcript information with targeted RNA-seq
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Home › Learning centers › Next-generation sequencing › Technical notes › RNA-seq › Stranded libraries from 100 ng - 1 ug total RNA

Technical notes

  • Single-cell RNA- and DNA-seq
    • All-in-one cDNA synthesis and library prep from single cells
    • Highest sensitivity for single-cell mRNA-seq
    • Stranded libraries from single cells
    • Streamlined single-cell mRNA-seq
    • Full-length mRNA libraries from single cells (SMART-Seq v4)
    • 3' mRNA libraries from single cells (SMART-Seq v4 3' DE Kit)
    • Full-length mRNA libraries from single cells for Fluidigm C1 (SMART-Seq v4)
    • Full-length single-cell library method comparison
    • High-resolution CNV detection using PicoPLEX Gold DNA-Seq
    • Next-gen WGA method for CNV and SNV detection from single cells
  • RNA-seq
    • Stranded libraries from picogram-input total RNA (v3)
    • Automation-friendly, all-in-one cDNA synthesis and library prep
    • All-in-one cDNA synthesis and library prep from ultra-low RNA inputs
    • Stranded libraries from picogram-input total RNA (v2)
    • Stranded libraries from FFPE inputs (v2)
    • Stranded libraries from 100 ng - 1 ug total RNA
    • Stranded libraries from 100 pg-100 ng total RNA
    • Stranded libraries from picogram-input total RNA (v1)
    • Stranded RNA-seq competitor kit comparison
    • Nonstranded libraries from FFPE inputs
    • Sensitive capture of full-length transcript information with targeted RNA-seq
  • DNA-seq
    • Comparing ThruPLEX HV PLUS to Kapa and NEBNext
    • Improvements to ThruPLEX HV
    • ThruPLEX HV outperforms NEBNext Ultra II
    • Accurate detection of low-frequency variants using molecular tags
    • Streamlined DNA-seq from challenging samples
    • Low cell number ChIP-seq using SMARTer ThruPLEX
    • Cell-free DNA sequencing
    • Sequencing analysis of low-frequency mutations in cfDNA
    • DNA-seq from FFPE samples
    • Low-input whole-exome sequencing
    • Tag-seq variant detection
    • Low-volume DNA shearing for SMARTer ThruPLEX library prep
  • Immune profiling
    • BCR repertoire profiling from human samples (bulk)
    • Improved TCR repertoire profiling from human samples (bulk)
    • TCR repertoire profiling from human samples (single cells)
    • TCR repertoire profiling from human samples (bulk)
    • TCR repertoire profiling from mouse samples (bulk)
    • BCR repertoire profiling from mouse samples (bulk)
  • Epigenetics and smRNA-seq
    • Full-length small RNA libraries
    • ChIP-seq libraries for transcription factor analysis
    • ChIP-seq libraries from ssDNA
    • Methylated DNA-seq
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Tech Note

A complete solution for generating stranded RNA-seq libraries from high-input total RNA

  • Fast, efficient rRNA depletion and stranded library preparation
  • Reproducibility and consistency across replicates
  • High-quality, stranded data
  • Accurate results regardless of RNA quality
Introduction Results Conclusions Methods References

Introduction  

Streamlined methods provide the opportunities necessary to push each experiment to its fullest potential. By combining time-saving techniques with high-performance reagents, studies in transcriptomics can move forward efficiently and accurately. Expression analysis of the entire transcriptome by RNA-sequencing (RNA-seq) can reap great benefits from highly sensitive, versatile, and easy-to-use protocols. Traditionally, generation of RNA-seq libraries from total RNA has been challenged by the high amounts of ribosomal RNA (rRNA) in the starting material, and lengthy protocols required to incorporate platform-specific adaptors via ligation. The SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian is a unique solution for generating indexed cDNA libraries suitable for next-generation sequencing (NGS) on any Illumina platform, starting with 100 ng–1 µg of total mammalian RNA of any quality.

Results  

Fast, accurate technology for rRNA removal and library generation

Our SMARTer RNA-seq kits are based on the core SMART (Switching Mechanism at 5' End of RNA Template) technology (Chenchik et al. 1998), a streamlined process that maintains strand information, and also eliminates tedious library preparation by incorporating adaptors in reverse transcription and PCR steps. The strand-specific reverse transcription reaction maintains close to 99% accurate strand-of-origin information, allowing for the identification of overlapping transcripts and antisense transcripts. Sequencing-ready libraries are generated during PCR amplification of the cDNA, using primers containing Illumina cluster-generating sequences and indexes.

Total RNA can consist of ≥90% rRNA, making it important to remove rRNA from samples before generating RNA-seq libraries. The RiboGone technology incorporated in the protocol uses hybridization technology and RNase H digestion to bind and specifically deplete nuclear rRNA sequences (5S, 5.8S, 18S, and 28S), as well as mitochondrial rRNA sequences (12S) from human, mouse, or rat total RNA (Morlan, Qu, and Sinicropi 2012). By depleting the rRNA in samples prior to library generation, sequencing costs are lowered and mapping statistics are improved.

With the combined power of SMART and RiboGone technologies, the SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian enables you to go from total RNA to Illumina-compatible RNA-seq libraries in around five hours.

Generate Illumina-specific, stranded RNA-seq libraries in 5 hours from total RNA

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation. Panel A. Depletion of rRNA from total RNA samples with RiboGone technology. Panel B. First-strand cDNA synthesis with SMART technology, incorporating Illumina Read Primers 1 and 2. Panel C. Template switching and generation of sequencing libraries with Illumina cluster-generating sequences and indexes by PCR amplification.

Reproducible sequencing data

The SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian produces extremely reliable RNA-seq data. Two 100-ng samples of Human Universal Reference RNA (HURR; Agilent) were treated with this kit, and the data from the two resulting libraries were compared. The high correlation between them (R = 0.99) displays an impressive level of reproducibility and consistency across replicates.

High correlation of RNA-seq data between replicates

Reproducibility across replicates. RNA-seq libraries were generated from two samples of 100 ng of HURR. The scatterplot illustrates correlations between the FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) from the two libraries.

High-quality sequencing data

With the SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian system, researchers can generate RNA-seq libraries from a variety of samples, including HURR and HBRR (Human Brain Reference RNA; Ambion), starting from 100 ng–1 µg of total RNA. Using this kit, rRNA content was depleted from samples prior to cDNA synthesis and library generation. When sequenced, both the HURR and HBRR libraries yielded a high number of quality reads, with 88–94% mapped, 84–91% uniquely mapped, and approximately 17,600 genes identified. Additionally, based on the ERCC Spike-In RNA, strand information was maintained at about 99% for both samples. The benefits of rRNA depletion are clear, with less than 0.5% of reads from the HURR library and less than 6% of reads from the HBRR library mapped to rRNA.

Sequence alignment metrics
RNA source Human Universal Human Brain
Input amount 400 ng
Number of reads (millions) 8.5 (paired end reads)
Percentage of reads (%):
rRNA 0.3% 5.3%
Mapped to genome 94% 88%
Mapped uniquely to genome 91% 84%
Exonic 43% 50%
Intronic 43% 33%
Intergenic 14% 12%
Number of genes identified 17,570 17,600
Percentage of ERCC transcripts with correct strand 99.3% 98.8%


Sequence Alignment Metrics.
 400 ng of HURR and HBRR with ERCC Spike-In RNA were treated with this kit. Alignment data is displayed for both libraries, with the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

These same RNA-seq libraries, generated from HURR and HBRR samples, produced data that had a strong correlation (R = 0.927) with qPCR data for the same RNAs obtained through the MicroArray Quality Control (MAQC) analysis. This suggested that the RiboGone method of rRNA depletion and SMARTer cDNA synthesis and library preparation did not negatively affect the RNA-seq data and maintained exceptional accuracy.

MAQC analysis shows high accuracy and strong correlation with pPCR data

MAQC Analysis. RNA-seq libraries were generated with 400 ng of HURR and HBRR. The scatter plot shows the Log2 ratio of FPKMs of HURR/HBRR graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes.

RNA-seq libraries produced with this kit provide an accurate representation of your sample. A 400-ng HBRR sample with ERCC (External RNA Controls Consortium) Spike-In RNA Mix (Life Technologies) was treated with this kit, and the libraries were sequenced, generating 8.5 million paired end reads. The FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) showed a strong correlation (R2 = 0.9199) and linearity (slope = 0.9988) to the input concentrations of the individual ERCC transcripts, indicating excellent accuracy and dynamic range.

ERCC spike-in data shows excellent accuracy and dynamic range

Dynamic range and linearity of RNA-seq data. Libraries were generated from Human Brain Reference RNA with ERCC Spike-In RNA Mix2. The above graph shows strong correlation between the Log2 of input concentrations of individual ERCC transcripts vs. the Log2 of FPKMs for those transcripts.

Reliable, accurate results across RNA inputs of varying quality

Libraries can be quickly and precisely generated from input RNA of a wide range of quality. Mouse Liver RNA was chemically sheared to a RIN (RNA Integrity Number) of either 3 or 7 (Mortazavi et al. 2008). Samples of each quality were used at both 100-ng and 1-µg levels and treated with this kit. All of the libraries generated from these samples had high mapping statistics with 81–88% mapped reads, 72–77% uniquely mapped reads, with over 12,000 genes identified. Stranded information of the biological RNA was maintained at high levels (95–98%), regardless of RIN value.

Sequence alignment metrics from RNA of varying quality
RNA source Mouse Liver
RNA quality (RIN) RIN 3 RIN 7
Input amount 100 ng 1 µg 100 ng 1 µg
Number of reads (millions) 1.7 (paired end reads)
Percentage of reads (%):
rRNA 2% 2% 1% 1%
Mapped to genome 82% 86% 81% 88%
Mapped uniquely to genome 73% 75% 72% 77%
Exonic 55% 53% 54% 54%
Intronic 32% 31% 33% 32%
Intergenic 12% 14% 12% 13%
Number of genes identified 12,079 12,172 12,099 12,212
Percent biological strandedness 95.5% 97.2% 95.6% 98.1%


High-quality libraries across varying levels of RNA quality.
 Libraries were generated from Mouse Liver RNA by chemical shearing until it had a RIN of 3 or 7. Sequencing data showed the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

The above data shows that the high reproducibility standards of this kit are not affected by the quality of input RNA. A comparison of data from the 1-µg libraries described above shows an extremely high correlation (R = 0.99), indicating the strong ability of the SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian kit to generate reliable, reproducible data across varying levels of RNA quality.

High quality, reproducible RNA-seq data from samples of varying RNA quality

Reproducibility across RNA quality. A scatterplot illustrates the correlations between the FPKMs from two libraries generated from 1 µg of Mouse Liver RNA that was chemically sheared until it had a RIN of 3 or 7.

Conclusions  

The SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian is a complete solution for preparing indexed Illumina sequencing libraries from 100 ng–1 µg of mammalian total RNA. This kit incorporates key RiboGone and SMART technologies, seamlessly blending abundant transcript (rRNA) removal and strand-specific library generation. SMART technology allows the addition of Illumina adaptors in a ligation-free manner, significantly reducing hands-on time while also increasing efficiency. The sequencing data obtained with this kit maintains high quality and reproducibility across sample replicates and RNA quality.

Methods  

Reproducibility across replicates:

Reproducibility across replicates was illustrated with two samples of 100 ng of Human Universal Reference RNA (Agilent), treated with SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian. The two replicates underwent the same protocol, except Replica #1 used 13 PCR cycles and Replica #2 used 14 PCR cycles. The libraries were sequenced at 1.3 million single end reads (1 x 50 bp) on an Illumina MiSeq® instrument, and aligned with STAR against hg19 with Ensembl annotation. 

MAQC and ERCC analysis:

The quality of sequencing data was demonstrated via MAQC analysis, dynamic range analysis, and sequence alignment metrics. For this purpose, RNA-seq libraries were generated from 400 ng of Human Universal Reference RNA (HURR; Agilent) and Human Brain Reference RNA (HBRR; Ambion) with ERCC Spike-In RNA, with Mix1 used for HURR and Mix2 used for HBRR. The libraries were sequenced at 8.5 million paired end reads (2 x 75 bp) on an Illumina MiSeq instrument, and aligned with STAR against hg19 with Ensembl annotation. The percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand were defined by Picard analysis. For MAQC analysis, the Log2 ratio of FPKMs from HURR/HBRR was graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes. For the dynamic range study, the Log2 of input concentrations of individual ERCC transcripts was graphed against the Log2 of FPKMs for those transcripts in the HBRR sample. 

Library generation across RNA inputs of varying quality

To compare data across RNA quality, Mouse Liver RNA was chemically sheared until it had a RIN (RNA Integrity Number) of 3 or 7. Either 100 ng or 1 µg of each RIN was used with this kit to generate RNA-seq libraries. These libraries were sequenced at 1.7 million paired end reads (2 x 25 bp) on an Illumina MiSeq instrument, and aligned with STAR against mm10 with Ensembl annotation. The percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand were defined by Picard analysis. Correlations between the FPKMs of libraries generated from 1 µg of RNA with both RINs were illustrated in a scatterplot.

References  

Chenchik, A. et al. Generation and use of high-quality cDNA from small amounts of total RNA by SMART PCR. Gene cloning Anal. by RT-PCR 305–319 (1998).

Morlan, J. D., Qu, K. & Sinicropi, D. V. Selective depletion of rRNA enables whole transcriptome profiling of archival fixed tissue. PLoS One 7, (2012). 

Mortazavi, A., Williams, B. A., McCue, K., Schaeffer, L. & Wold, B. Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat. Methods 5, 621–628 (2008).     

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The SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian generates libraries for RNA sequencing (RNA-seq), compatible with Illumina platforms for mammalian samples. The kit is designed to work with input ranges from 100 ng to 1 μg of total RNA. This kit incorporates both RiboGone and SMART technology, and processes ribosomal RNA (rRNA) removal (5S, 5.8S, 18S, and 28S nuclear rRNA sequences), as well as some mitochondrial rRNA sequences (12S), from human, mouse, or rat full-length or sheared total RNA, followed by cDNA synthesis and Illumina library preparation. RNA-seq libraries prepared with this kit are indexed.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Reproducibility across replicates

Reproducibility across replicates
Reproducibility across replicates. RNA-seq libraries were generated from two samples of 100 ng of HURR. The scatterplot illustrates correlations between the FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) from the two libraries.

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Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation
Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation. Section A. Depletion of rRNA from total RNA samples with RiboGone technology. Section B. First-strand cDNA synthesis with SMART technology, incorporating Illumina Read Primers 1 and 2. Section C. Template switching and generation of sequencing libraries with Illumina cluster-generating sequences and indexes by PCR amplification

Back

MAQC Analysis

MAQC Analysis
MAQC Analysis. RNA-seq libraries were generated with 400 ng of HURR and HUBR. The scatter plot shows the Log2 ratio of FPKMs of HURR/HBRR graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes.

Back

Dynamic range and linearity of RNA-seq data

Dynamic range and linearity of RNA-seq data
Dynamic range and linearity of RNA-seq data. Libraries were generated from Human Brain Reference RNA with ERCC Spike-In RNA Mix2. The above graph shows strong correlation between the Log2 of input concentrations of individual ERCC transcripts vs. the Log2 of FPKMs for those transcripts.

Back

Reproducibility across RNA quality

Reproducibility across RNA quality
Reproducibility across RNA quality. A scatterplot illustrates the correlations between the FPKMs from two libraries generated from 1 µg Mouse Liver RNA that was chemically sheared until it had a RIN of 3 or 7.

Back

Sequence Alignment Metrics

Sequence Alignment Metrics
Sequence Alignment Metrics. 400 ng of HURR and HBRR with ERCC Spike-In RNA were treated with this kit. Alignment data is displayed for both libraries, with the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

Back

High-quality libraries across varying levels of RNA quality

High-quality libraries across varying levels of RNA quality
High-quality libraries across varying levels of RNA quality. Libraries were generated from Mouse Liver RNA by chemically shearing until it had a RIN of 3 or 7. Sequencing data showed the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.
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Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Back

Reproducibility across replicates

Reproducibility across replicates
Reproducibility across replicates. RNA-seq libraries were generated from two samples of 100 ng of HURR. The scatterplot illustrates correlations between the FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) from the two libraries.

Back

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation
Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation. Section A. Depletion of rRNA from total RNA samples with RiboGone technology. Section B. First-strand cDNA synthesis with SMART technology, incorporating Illumina Read Primers 1 and 2. Section C. Template switching and generation of sequencing libraries with Illumina cluster-generating sequences and indexes by PCR amplification

Back

MAQC Analysis

MAQC Analysis
MAQC Analysis. RNA-seq libraries were generated with 400 ng of HURR and HUBR. The scatter plot shows the Log2 ratio of FPKMs of HURR/HBRR graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes.

Back

Dynamic range and linearity of RNA-seq data

Dynamic range and linearity of RNA-seq data
Dynamic range and linearity of RNA-seq data. Libraries were generated from Human Brain Reference RNA with ERCC Spike-In RNA Mix2. The above graph shows strong correlation between the Log2 of input concentrations of individual ERCC transcripts vs. the Log2 of FPKMs for those transcripts.

Back

Reproducibility across RNA quality

Reproducibility across RNA quality
Reproducibility across RNA quality. A scatterplot illustrates the correlations between the FPKMs from two libraries generated from 1 µg Mouse Liver RNA that was chemically sheared until it had a RIN of 3 or 7.

Back

Sequence Alignment Metrics

Sequence Alignment Metrics
Sequence Alignment Metrics. 400 ng of HURR and HBRR with ERCC Spike-In RNA were treated with this kit. Alignment data is displayed for both libraries, with the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

Back

High-quality libraries across varying levels of RNA quality

High-quality libraries across varying levels of RNA quality
High-quality libraries across varying levels of RNA quality. Libraries were generated from Mouse Liver RNA by chemically shearing until it had a RIN of 3 or 7. Sequencing data showed the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.
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Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Back

Reproducibility across replicates

Reproducibility across replicates
Reproducibility across replicates. RNA-seq libraries were generated from two samples of 100 ng of HURR. The scatterplot illustrates correlations between the FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) from the two libraries.

Back

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation
Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation. Section A. Depletion of rRNA from total RNA samples with RiboGone technology. Section B. First-strand cDNA synthesis with SMART technology, incorporating Illumina Read Primers 1 and 2. Section C. Template switching and generation of sequencing libraries with Illumina cluster-generating sequences and indexes by PCR amplification

Back

MAQC Analysis

MAQC Analysis
MAQC Analysis. RNA-seq libraries were generated with 400 ng of HURR and HUBR. The scatter plot shows the Log2 ratio of FPKMs of HURR/HBRR graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes.

Back

Dynamic range and linearity of RNA-seq data

Dynamic range and linearity of RNA-seq data
Dynamic range and linearity of RNA-seq data. Libraries were generated from Human Brain Reference RNA with ERCC Spike-In RNA Mix2. The above graph shows strong correlation between the Log2 of input concentrations of individual ERCC transcripts vs. the Log2 of FPKMs for those transcripts.

Back

Reproducibility across RNA quality

Reproducibility across RNA quality
Reproducibility across RNA quality. A scatterplot illustrates the correlations between the FPKMs from two libraries generated from 1 µg Mouse Liver RNA that was chemically sheared until it had a RIN of 3 or 7.

Back

Sequence Alignment Metrics

Sequence Alignment Metrics
Sequence Alignment Metrics. 400 ng of HURR and HBRR with ERCC Spike-In RNA were treated with this kit. Alignment data is displayed for both libraries, with the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

Back

High-quality libraries across varying levels of RNA quality

High-quality libraries across varying levels of RNA quality
High-quality libraries across varying levels of RNA quality. Libraries were generated from Mouse Liver RNA by chemically shearing until it had a RIN of 3 or 7. Sequencing data showed the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.
634876 SMARTer® Stranded Total RNA Sample Prep Kit - HI Mammalian 96 Rxns USD $4977.00

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Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Documents Components You May Also Like Image Data Resources

Back

Reproducibility across replicates

Reproducibility across replicates
Reproducibility across replicates. RNA-seq libraries were generated from two samples of 100 ng of HURR. The scatterplot illustrates correlations between the FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) from the two libraries.

Back

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation
Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation. Section A. Depletion of rRNA from total RNA samples with RiboGone technology. Section B. First-strand cDNA synthesis with SMART technology, incorporating Illumina Read Primers 1 and 2. Section C. Template switching and generation of sequencing libraries with Illumina cluster-generating sequences and indexes by PCR amplification

Back

MAQC Analysis

MAQC Analysis
MAQC Analysis. RNA-seq libraries were generated with 400 ng of HURR and HUBR. The scatter plot shows the Log2 ratio of FPKMs of HURR/HBRR graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes.

Back

Dynamic range and linearity of RNA-seq data

Dynamic range and linearity of RNA-seq data
Dynamic range and linearity of RNA-seq data. Libraries were generated from Human Brain Reference RNA with ERCC Spike-In RNA Mix2. The above graph shows strong correlation between the Log2 of input concentrations of individual ERCC transcripts vs. the Log2 of FPKMs for those transcripts.

Back

Reproducibility across RNA quality

Reproducibility across RNA quality
Reproducibility across RNA quality. A scatterplot illustrates the correlations between the FPKMs from two libraries generated from 1 µg Mouse Liver RNA that was chemically sheared until it had a RIN of 3 or 7.

Back

Sequence Alignment Metrics

Sequence Alignment Metrics
Sequence Alignment Metrics. 400 ng of HURR and HBRR with ERCC Spike-In RNA were treated with this kit. Alignment data is displayed for both libraries, with the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

Back

High-quality libraries across varying levels of RNA quality

High-quality libraries across varying levels of RNA quality
High-quality libraries across varying levels of RNA quality. Libraries were generated from Mouse Liver RNA by chemically shearing until it had a RIN of 3 or 7. Sequencing data showed the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.
634877 SMARTer® Stranded Total RNA Sample Prep Kit - HI Mammalian 192 Rxns USD $8900.00

The SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian generates libraries for RNA sequencing (RNA-seq), compatible with Illumina platforms for mammalian samples. The kit is designed to work with input ranges from 100 ng to 1 μg of total RNA. This kit incorporates both RiboGone and SMART technology, and processes ribosomal RNA (rRNA) removal (5S, 5.8S, 18S, and 28S nuclear rRNA sequences), as well as some mitochondrial rRNA sequences (12S), from human, mouse, or rat full-length or sheared total RNA, followed by cDNA synthesis and Illumina library preparation. RNA-seq libraries prepared with this kit are indexed.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Documents Components You May Also Like Image Data Resources

Back

Reproducibility across replicates

Reproducibility across replicates
Reproducibility across replicates. RNA-seq libraries were generated from two samples of 100 ng of HURR. The scatterplot illustrates correlations between the FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) from the two libraries.

Back

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation
Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation. Section A. Depletion of rRNA from total RNA samples with RiboGone technology. Section B. First-strand cDNA synthesis with SMART technology, incorporating Illumina Read Primers 1 and 2. Section C. Template switching and generation of sequencing libraries with Illumina cluster-generating sequences and indexes by PCR amplification

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MAQC Analysis

MAQC Analysis
MAQC Analysis. RNA-seq libraries were generated with 400 ng of HURR and HUBR. The scatter plot shows the Log2 ratio of FPKMs of HURR/HBRR graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes.

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Dynamic range and linearity of RNA-seq data

Dynamic range and linearity of RNA-seq data
Dynamic range and linearity of RNA-seq data. Libraries were generated from Human Brain Reference RNA with ERCC Spike-In RNA Mix2. The above graph shows strong correlation between the Log2 of input concentrations of individual ERCC transcripts vs. the Log2 of FPKMs for those transcripts.

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Reproducibility across RNA quality

Reproducibility across RNA quality
Reproducibility across RNA quality. A scatterplot illustrates the correlations between the FPKMs from two libraries generated from 1 µg Mouse Liver RNA that was chemically sheared until it had a RIN of 3 or 7.

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Sequence Alignment Metrics

Sequence Alignment Metrics
Sequence Alignment Metrics. 400 ng of HURR and HBRR with ERCC Spike-In RNA were treated with this kit. Alignment data is displayed for both libraries, with the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

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High-quality libraries across varying levels of RNA quality

High-quality libraries across varying levels of RNA quality
High-quality libraries across varying levels of RNA quality. Libraries were generated from Mouse Liver RNA by chemically shearing until it had a RIN of 3 or 7. Sequencing data showed the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.
634878 SMARTer® Stranded Total RNA Sample Prep Kit - HI Mammalian 480 Rxns USD $18254.00

The SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian generates libraries for RNA sequencing (RNA-seq), compatible with Illumina platforms for mammalian samples. The kit is designed to work with input ranges from 100 ng to 1 μg of total RNA. This kit incorporates both RiboGone and SMART technology, and processes ribosomal RNA (rRNA) removal (5S, 5.8S, 18S, and 28S nuclear rRNA sequences), as well as some mitochondrial rRNA sequences (12S), from human, mouse, or rat full-length or sheared total RNA, followed by cDNA synthesis and Illumina library preparation. RNA-seq libraries prepared with this kit are indexed.

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Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Reproducibility across replicates

Reproducibility across replicates
Reproducibility across replicates. RNA-seq libraries were generated from two samples of 100 ng of HURR. The scatterplot illustrates correlations between the FPKMs (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) from the two libraries.

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Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation

Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation
Flowchart of SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian library generation. Section A. Depletion of rRNA from total RNA samples with RiboGone technology. Section B. First-strand cDNA synthesis with SMART technology, incorporating Illumina Read Primers 1 and 2. Section C. Template switching and generation of sequencing libraries with Illumina cluster-generating sequences and indexes by PCR amplification

Back

MAQC Analysis

MAQC Analysis
MAQC Analysis. RNA-seq libraries were generated with 400 ng of HURR and HUBR. The scatter plot shows the Log2 ratio of FPKMs of HURR/HBRR graphed against the Log2 of the ratio of HURR/HBRR derived from qPCR Taqman probes.

Back

Dynamic range and linearity of RNA-seq data

Dynamic range and linearity of RNA-seq data
Dynamic range and linearity of RNA-seq data. Libraries were generated from Human Brain Reference RNA with ERCC Spike-In RNA Mix2. The above graph shows strong correlation between the Log2 of input concentrations of individual ERCC transcripts vs. the Log2 of FPKMs for those transcripts.

Back

Reproducibility across RNA quality

Reproducibility across RNA quality
Reproducibility across RNA quality. A scatterplot illustrates the correlations between the FPKMs from two libraries generated from 1 µg Mouse Liver RNA that was chemically sheared until it had a RIN of 3 or 7.

Back

Sequence Alignment Metrics

Sequence Alignment Metrics
Sequence Alignment Metrics. 400 ng of HURR and HBRR with ERCC Spike-In RNA were treated with this kit. Alignment data is displayed for both libraries, with the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

Back

High-quality libraries across varying levels of RNA quality

High-quality libraries across varying levels of RNA quality
High-quality libraries across varying levels of RNA quality. Libraries were generated from Mouse Liver RNA by chemically shearing until it had a RIN of 3 or 7. Sequencing data showed the percentage of reads that mapped to rRNA, exonic regions, intronic regions, intergenic regions, and the correct strand, as defined by Picard analysis.

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