ArticleProceedings of the National Academy of Sciences of the United States of America2014
Molecular indexing enables quantitative targeted RNA sequencing and reveals poor efficiencies in standard library preparations.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 62 papers.
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Who cites it
62 citing papers in PubMed, 115 citations in OpenAlex.
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- Loosening the Lid on Shoulder Osteoarthritis: How the Transcriptome and Metabolic Syndrome Correlate with End-Stage Disease.International journal of molecular sciences · 2025Article
- Chromato-kinetic fingerprinting enables multiomic digital counting of single disease biomarker molecules.bioRxiv : the preprint server for biology · 2025Article
- Subregion-specific transcriptomic profiling of rat brain reveals sex-distinct gene expression impacted by adolescent stress.Neuroscience · 2024Article
- Genomic reproducibility in the bioinformatics era.Genome biology · 2024Review
- Digital RNA sequencing using unique molecular identifiers enables ultrasensitive RNA mutation analysis.Communications biology · 2024Article
- Methodologies for bacterial ribonuclease characterization using RNA-seq.FEMS microbiology reviews · 2023Review
- Synergising single-cell resolution and 4sU labelling boosts inference of transcriptional bursting.Genome biology · 2023Article
- Transcriptomics for Clinical and Experimental Biology Research: Hang on a Seq.Advanced genetics (Hoboken, N.J.) · 2023Review
- The human pathobiontProceedings of the National Academy of Sciences of the United States of America · 2022Article
- DeLTa-Seq: direct-lysate targeted RNA-Seq from crude tissue lysate.Plant methods · 2022Article
- Transcriptional Landscape of Cotton Fiber Development and Its Alliance With Fiber-Associated Traits.Frontiers in plant science · 2022Article
- FDA-led consortium studies advance quality control of targeted next generation sequencing assays for precision oncology.Precision cancer medicine · 2021Article
- Advancing NGS quality control to enable measurement of actionable mutations in circulating tumor DNA.Cell reports methods · 2021Article
- The role of noncoding RNAs in Parkinson's disease: biomarkers and associations with pathogenic pathways.Journal of biomedical science · 2021Review
- Development of a program for in silico optimized selection of oligonucleotide-based molecular barcodes.PloS one · 2021Article
- Integration of mRNA and miRNA Analysis Reveals the Molecular Mechanism of Cotton Response to Salt Stress.Frontiers in plant science · 2021Article
- Epigenetic Regulation of the Vascular Endothelium by Angiogenic LncRNAs.Frontiers in genetics · 2021Review
2 more citing papers are in PubMed but not listed here.
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Authors and funding
7 authors at 2 institutions in 1 country.
Funding
Abstract
We present a simple molecular indexing method for quantitative targeted RNA sequencing, in which mRNAs of interest are selectively captured from complex cDNA libraries and sequenced to determine their absolute concentrations. cDNA fragments are individually labeled so that each molecule can be tracked from the original sample through the library preparation and sequencing process. Multiple copies of cDNA fragments of identical sequence become distinct through labeling, and replicate clones created during PCR amplification steps can be identified and assigned to their distinct parent molecules. Selective capture enables efficient use of sequencing for deep sampling and for the absolute quantitation of rare or transient transcripts that would otherwise escape detection by standard sequencing methods. We have also constructed a set of synthetic barcoded RNA molecules, which can be introduced as controls into the sample preparation mix and used to monitor the efficiency of library construction. The quantitative targeted sequencing revealed extremely low efficiency in standard library preparations, which were further confirmed by using synthetic barcoded RNA molecules. This finding shows that standard library preparation methods result in the loss of rare transcripts and highlights the need for monitoring library efficiency and for developing more efficient sample preparation methods.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.