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Related Concept Videos

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The atomic mass of an element varies due to the relative ratio of its isotopes. A sample's relative proportion of oxygen isotopes influences its average atomic mass. For instance, if we were to measure the atomic mass of oxygen from a sample, the mass would be a weighted average of the isotopic masses of oxygen in that sample. Since a single sample is not likely to perfectly reflect the true atomic mass of oxygen for all the molecules of oxygen on Earth, the mass we obtain from this...
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Intact DNA strands can be found in fossils, while scientists sometimes struggle to keep RNA intact under laboratory conditions. The structural variations between RNA and DNA underlie the differences in their stability and longevity. Because DNA is double-stranded, it is inherently more stable. The single-stranded structure of RNA is less stable but also more flexible and can form weak internal bonds. Additionally, most RNAs in the cell are relatively short, while DNA can be up to 250 million...
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RIBO-seq in Bacteria: a Sample Collection and Library Preparation Protocol for NGS Sequencing
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Systematic evaluation of RNA-Seq preparation protocol performance.

Hsueh-Ping Chao1,2, Yueping Chen1, Yoko Takata1

  • 1Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Science Park, Smithville, TX, 78957, USA.

BMC Genomics
|July 13, 2019
PubMed
Summary
This summary is machine-generated.

Choosing the right RNA-Seq library preparation kit is crucial for accurate gene expression analysis. This study evaluates four kits, finding that while all distinguish experimental groups, kit selection impacts gene recovery and data outcomes.

Keywords:
Next generation sequencingQuality controlRNA-Seq

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Identification of Footprints of RNA:Protein Complexes via RNA Immunoprecipitation in Tandem Followed by Sequencing RIPiT-Seq
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Area of Science:

  • Molecular Biology
  • Genomics
  • Bioinformatics

Background:

  • RNA-Sequencing (RNA-Seq) is a key tool for whole-transcriptome analysis.
  • Numerous commercial kits exist for RNA-Seq library preparation, but their performance variations are not fully understood.
  • Understanding kit limitations is vital due to the cost and time investment in RNA-Seq.

Purpose of the Study:

  • To comprehensively evaluate and compare the performance of four commonly used RNA-Seq library preparation kits.
  • To assess kit performance regarding ribosomal RNA removal, read coverage, gene expression profiling, and non-coding RNA detection.
  • To provide insights into selecting the optimal RNA-Seq kit for specific research objectives.

Main Methods:

  • Evaluation of four RNA-Seq kits: Illumina TruSeq Stranded Total RNA, Illumina TruSeq Stranded mRNA, modified NuGEN Ovation v2, and TaKaRa SMARTer Ultra Low RNA Kit v3.
  • Assessment of key performance metrics including reproducibility, 5'/3' end bias, differentially expressed genes (DEGs), long non-coding RNAs (lncRNAs), and alternatively spliced transcripts.
  • Analysis of RNA input levels and their impact on kit performance.

Main Results:

  • All evaluated kits were suitable for distinguishing between experimental groups at recommended RNA input levels.
  • Illumina kits showed similarity in DEG recovery; Illumina, NuGEN, and TaKaRa kits identified overlapping DEGs but also enriched distinct gene sets.
  • TruSeq protocols favored genes with higher expression and GC content, while the NuGEN kit captured longer genes.

Conclusions:

  • The choice of RNA-Seq library preparation kit significantly influences experimental data outcomes.
  • The TruSeq Stranded mRNA kit is versatile for protein-coding gene studies.
  • The TaKaRa SMARTer Ultra Low RNA Kit may be suitable for low RNA input, but the TruSeq mRNA kit outperformed it at standard input levels for rRNA removal and DEG recovery.