SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian
SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian
Brand: Takara Bio.
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SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian
Strand-specific RNA-seq libraries for Illumina platforms from high-input total RNA
The SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian provides a rapid solution for preparing indexed Illumina sequencing libraries from 100 ng–1 µg of mammalian total RNA. This kit combines RiboGone and SMART technologies, seamlessly incorporating efficient rRNA removal followed by strand-specific library generation in around 5 hours. Because rRNA can comprise ~90% or more of total RNA, effective depletion of rRNA prior to library generation can lower sequencing costs and improve mapping statistics. SMART cDNA synthesis technology followed by PCR complete a streamlined process that performs strand information and eliminates downstream library prep by incorporating indexes and adapters during the reverse transcription and amplification steps.
These high-input kits contain 12 reverse primers and up to 8 forward primers that allow for multiplexing of up to 96 separate libraries in a high-throughput format. The specific primers you receive will depend on your particular kit. The entire protocol—from rRNA depletion through creation of Illumina-specific, indexed RNA-seq libraries—takes only about 5 hours.
Overview
- Strand information—Identify each transcript’s strand of origin with >99% accuracy.
- Fast, streamlined protocol—Go from start to finish in ~5 hours.
- Broad input range—Use 100 ng–1 µg of total RNA from human, mouse, or rat.
- Versatility—Get reliable data across replicates. See high-quality, reproducible data from total RNA with a RIN (RNA Integrity Number) from 3–10.
- Seamless integration with Illumina sequencing—Generate up to 192 uniquely indexed Illumina-ready libraries.
Applications
- RNA-seq for mammalian samples on Illumina platforms
- NGS library generation that retains strand information
- Analysis of coding and noncoding sequence information
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.
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
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.
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.
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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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.
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
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.
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.
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.
