Dataset Information

TM3'seq: A Tagmentation-Mediated 3' Sequencing Approach for Improving Scalability of RNAseq Experiments.

ABSTRACT: RNA-seq has become the standard tool for collecting genome-wide expression data in diverse fields, from quantitative genetics and medical genomics to ecology and developmental biology. However, RNA-seq library preparation is still prohibitive for many laboratories. Recently, the field of single-cell transcriptomics has reduced costs and increased throughput by adopting early barcoding and pooling of individual samples -producing a single final library containing all samples. In contrast, RNA-seq protocols where each sample is processed individually are significantly more expensive and lower throughput than single-cell approaches. Yet, many projects depend on individual library generation to preserve important samples or for follow-up re-sequencing experiments. Improving on currently available RNA-seq methods we have developed TM3'seq, a 3'-enriched library preparation protocol that uses Tn5 transposase and preserves sample identity at each step. TM3'seq is designed for high-throughput processing of individual samples (96 samples in 6h, with only 3h hands-on time) at a fraction of the cost of commercial kits ($1.5 per sample). The protocol was tested in a range of human and Drosophila melanogaster RNA samples, recovering transcriptomes of the same quality and reliability than the commercial NEBNext kit. We expect that the cost- and time-efficient features of TM3'seq make large-scale RNA-seq experiments more permissive for the entire scientific community.

SUBMITTER: Pallares LF

PROVIDER: S-EPMC6945013 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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TM3'seq: A Tagmentation-Mediated 3' Sequencing Approach for Improving Scalability of RNAseq Experiments.

Pallares Luisa F LF Picard Serge S Ayroles Julien F JF

G3 (Bethesda, Md.) 20200107 1

RNA-seq has become the standard tool for collecting genome-wide expression data in diverse fields, from quantitative genetics and medical genomics to ecology and developmental biology. However, RNA-seq library preparation is still prohibitive for many laboratories. Recently, the field of single-cell transcriptomics has reduced costs and increased throughput by adopting early barcoding and pooling of individual samples -producing a single final library containing all samples. In contrast, RNA-seq ...[more]

PMID: 31676507

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Project description:IntroductionLack of access to empirically-supported psychological treatments (EPT) that are contextually appropriate and feasible to deliver by non-specialist health workers (referred to as 'counsellors') are major barrier for the treatment of mental health problems in resource poor countries. To address this barrier, the 'Program for Effective Mental Health Interventions in Under-resourced Health Systems' (PREMIUM) designed a method for the development of EPT for severe depression and harmful drinking. This was implemented over three years in India. This study assessed the relative usefulness and costs of the five 'steps' (Systematic reviews, In-depth interviews, Key informant surveys, Workshops with international experts, and Workshops with local experts) in the first phase of identifying the strategies and theoretical model of the treatment and two 'steps' (Case series with specialists, and Case series and pilot trial with counsellors) in the second phase of enhancing the acceptability and feasibility of its delivery by counsellors in PREMIUM with the aim of arriving at a parsimonious set of steps for future investigators to use for developing scalable EPT.Data and methodsThe study used two sources of data: the usefulness ratings by the investigators and the resource utilization. The usefulness of each of the seven steps was assessed through the ratings by the investigators involved in the development of each of the two EPT, viz. Healthy Activity Program for severe depression and Counselling for Alcohol Problems for harmful drinking. Quantitative responses were elicited to rate the utility (usefulness/influence), followed by open-ended questions for explaining the rankings. The resources used by PREMIUM were computed in terms of time (months) and monetary costs.ResultsThe theoretical core of the new treatments were consistent with those of EPT derived from global evidence, viz. Behavioural Activation and Motivational Enhancement for severe depression and harmful drinking respectively, indicating the universal applicability of these theories. All the steps of both phases in PREMIUM contributed to the development of the final EPT. However, if there were significant resource constraints, the steps can be limited to workshops with international and local experts in the first phase, and case series with specialists, and case series and pilot trial with counsellors in the second phase.ConclusionsIntegrating global evidence with local knowledge and practices are complementary and feasible goals to contribute to the development of contextually appropriate and feasible EPT in resource poor country settings. The emerging EPT share similar theoretical frameworks to those described in the global evidence. The PREMIUM method has relevance for any setting where populations have poor access to EPT as its explicit goal is to develop scalable treatments.

Dataset Information

TM3'seq: A Tagmentation-Mediated 3' Sequencing Approach for Improving Scalability of RNAseq Experiments.

Publications

TM3'seq: A Tagmentation-Mediated 3' Sequencing Approach for Improving Scalability of RNAseq Experiments.

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