We are analyzing https://link.springer.com/article/10.1186/s12864-017-3827-y.

Title:
A comprehensive assessment of RNA-seq protocols for degraded and low-quantity samples | BMC Genomics
Description:
RNA-sequencing (RNA-seq) has emerged as one of the most sensitive tool for gene expression analysis. Among the library preparation methods available, the standard poly(A) + enrichment provides a comprehensive, detailed, and accurate view of polyadenylated RNAs. However, on samples of suboptimal quality ribosomal RNA depletion and exon capture methods have recently been reported as better alternatives. We compared for the first time three commercial Illumina library preparation kits (TruSeq Stranded mRNA, TruSeq Ribo-Zero rRNA Removal, and TruSeq RNA Access) as representatives of these three different approaches using well-established human reference RNA samples from the MAQC/SEQC consortium on a wide range of input amounts (from 100 ng down to 1 ng) and degradation levels (intact, degraded, and highly degraded). We assessed the accuracy of the generated expression values by comparison to gold standard TaqMan qPCR measurements and gained unprecedented insight into the limits of applicability in terms of input quantity and sample quality of each protocol. We found that each protocol generates highly reproducible results (R 2 > 0.92) on intact RNA samples down to input amounts of 10 ng. For degraded RNA samples, Ribo-Zero showed clear performance advantages over the other two protocols as it generated more accurate and better reproducible gene expression results even at very low input amounts such as 1 ng and 2 ng. For highly degraded RNA samples, RNA Access performed best generating reliable data down to 5 ng input. We found that the ribosomal RNA depletion protocol from Illumina works very well at amounts far below recommendation and over a good range of intact and degraded material. We also infer that the exome-capture protocol (RNA Access, Illumina) performs better than other methods on highly degraded and low amount samples.
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Keywords {🔍}

rna, samples, input, degraded, access, ribozero, protocols, amounts, truseq, article, pubmed, values, protocol, sample, google, scholar, rnaseq, sequencing, quality, gene, intact, data, genes, expression, low, fig, amount, reads, library, highly, cas, detected, rnas, fold, central, degradation, seqca, seqcb, log, number, preparation, methods, human, coding, kit, size, illumina, results, libraries, approach,

Topics {✒️}

marc sultan & guglielmo roma full-length mrna-seq enriched strand-specific libraries vgf-derived peptide tlqp-21 related subjects van keuren-jensen kr article download pdf detecting differential expression high-throughput rna sequencing raw rna-sequencing reads rna-seq libraries prior taqman qrt-pcr data open data policies oligo-dt coated beads exome capture rna-seq degraded samples-size peak full size image rna-seq transcriptome profiles log fold-change values truseq stranded mrna allele specific expression large-scale pcr step library preparation steps single-cell levels generate high-quality data pre-analytical factors breast cancer reveals privacy choices/manage cookies gene expression omnibus detecting gene expression library preparation protocol library preparation methods generate high-quality reads ribosomal rna depletion marc sultan gene expression analysis low quality rna rna access kit library prep approaches clinical cancer sequencing creative commons license translating rna sequencing quantify gene expression total rna sequencing gene type category taqman qpcr measurements quality levels tested low-quantity samples breast cancer progression abundant ribosomal rnas

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For instance, how does the newly available RNA Access protocol perform on degraded samples?

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WebPage:
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         headline:A comprehensive assessment of RNA-seq protocols for degraded and low-quantity samples
         description:RNA-sequencing (RNA-seq) has emerged as one of the most sensitive tool for gene expression analysis. Among the library preparation methods available, the standard poly(A) + enrichment provides a comprehensive, detailed, and accurate view of polyadenylated RNAs. However, on samples of suboptimal quality ribosomal RNA depletion and exon capture methods have recently been reported as better alternatives. We compared for the first time three commercial Illumina library preparation kits (TruSeq Stranded mRNA, TruSeq Ribo-Zero rRNA Removal, and TruSeq RNA Access) as representatives of these three different approaches using well-established human reference RNA samples from the MAQC/SEQC consortium on a wide range of input amounts (from 100 ng down to 1 ng) and degradation levels (intact, degraded, and highly degraded). We assessed the accuracy of the generated expression values by comparison to gold standard TaqMan qPCR measurements and gained unprecedented insight into the limits of applicability in terms of input quantity and sample quality of each protocol. We found that each protocol generates highly reproducible results (R 2 > 0.92) on intact RNA samples down to input amounts of 10 ng. For degraded RNA samples, Ribo-Zero showed clear performance advantages over the other two protocols as it generated more accurate and better reproducible gene expression results even at very low input amounts such as 1 ng and 2 ng. For highly degraded RNA samples, RNA Access performed best generating reliable data down to 5 ng input. We found that the ribosomal RNA depletion protocol from Illumina works very well at amounts far below recommendation and over a good range of intact and degraded material. We also infer that the exome-capture protocol (RNA Access, Illumina) performs better than other methods on highly degraded and low amount samples.
         datePublished:2017-06-05T00:00:00Z
         dateModified:2017-06-05T00:00:00Z
         pageStart:1
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            RNA-sequencing
            Expression profiling
            Benchmarking
            Low quality
            Low quantity
            Differential expression
            Life Sciences
            general
            Microarrays
            Proteomics
            Animal Genetics and Genomics
            Microbial Genetics and Genomics
            Plant Genetics and Genomics
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      headline:A comprehensive assessment of RNA-seq protocols for degraded and low-quantity samples
      description:RNA-sequencing (RNA-seq) has emerged as one of the most sensitive tool for gene expression analysis. Among the library preparation methods available, the standard poly(A) + enrichment provides a comprehensive, detailed, and accurate view of polyadenylated RNAs. However, on samples of suboptimal quality ribosomal RNA depletion and exon capture methods have recently been reported as better alternatives. We compared for the first time three commercial Illumina library preparation kits (TruSeq Stranded mRNA, TruSeq Ribo-Zero rRNA Removal, and TruSeq RNA Access) as representatives of these three different approaches using well-established human reference RNA samples from the MAQC/SEQC consortium on a wide range of input amounts (from 100 ng down to 1 ng) and degradation levels (intact, degraded, and highly degraded). We assessed the accuracy of the generated expression values by comparison to gold standard TaqMan qPCR measurements and gained unprecedented insight into the limits of applicability in terms of input quantity and sample quality of each protocol. We found that each protocol generates highly reproducible results (R 2 > 0.92) on intact RNA samples down to input amounts of 10 ng. For degraded RNA samples, Ribo-Zero showed clear performance advantages over the other two protocols as it generated more accurate and better reproducible gene expression results even at very low input amounts such as 1 ng and 2 ng. For highly degraded RNA samples, RNA Access performed best generating reliable data down to 5 ng input. We found that the ribosomal RNA depletion protocol from Illumina works very well at amounts far below recommendation and over a good range of intact and degraded material. We also infer that the exome-capture protocol (RNA Access, Illumina) performs better than other methods on highly degraded and low amount samples.
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      dateModified:2017-06-05T00:00:00Z
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         RNA-sequencing
         Expression profiling
         Benchmarking
         Low quality
         Low quantity
         Differential expression
         Life Sciences
         general
         Microarrays
         Proteomics
         Animal Genetics and Genomics
         Microbial Genetics and Genomics
         Plant Genetics and Genomics
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            affiliation:
                  name:Novartis Pharma AG
                  address:
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            type:Person
            name:Anita Fernandez
            affiliation:
                  name:Novartis Pharma AG
                  address:
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                     type:PostalAddress
                  type:Organization
            type:Person
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            affiliation:
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                  address:
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                     type:PostalAddress
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               type:PostalAddress
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            name:Novartis Pharma AG
            address:
               name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
               type:PostalAddress
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      name:Anita Fernandez
      affiliation:
            name:Novartis Pharma AG
            address:
               name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
               type:PostalAddress
            type:Organization
      name:Marc Sultan
      affiliation:
            name:Novartis Pharma AG
            address:
               name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
               type:PostalAddress
            type:Organization
      email:[email protected]
      name:Guglielmo Roma
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      name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
      name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
      name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
      name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
      name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland
      name:Novartis Institutes for Biomedical Research, Novartis Pharma AG, Basel, Switzerland

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