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Reference Long and Short-Read Transcriptomes of Human Peripheral Blood Lymphocytes

ABSTRACT: This SuperSeries is composed of the SubSeries listed below.

ORGANISM(S): Homo sapiens

PROVIDER: GSE202329 | GEO | 2022/09/16

REPOSITORIES: GEO

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Reference Long and Short-Read Transcriptomes of Human Peripheral Blood Lymphocytes

Project description:Reference Long and Short-Read Transcriptomes of Human Peripheral Blood Lymphocytes

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Reference long-read isoform-aware transcriptomes of 4 human peripheral blood lymphocyte subsets.

Project description:Long-read sequencing technologies such as isoform sequencing can generate highly accurate sequences of full-length mRNA transcript isoforms. Such long-read transcriptomics may be especially useful in investigations of lymphocyte functional plasticity as it relates to human health and disease. However, no long-read isoform-aware reference transcriptomes of human circulating lymphocytes are readily available despite being valuable as benchmarks in a variety of transcriptomic studies. To begin to fill this gap, we purified 4 lymphocyte populations (CD4+ T, CD8+ T, NK, and Pan B cells) from the peripheral blood of a healthy male donor and obtained high-quality RNA (RIN > 8) for isoform sequencing and parallel RNA-Seq analyses. Many novel polyadenylated transcript isoforms, supported by both isoform sequencing and RNA-Seq data, were identified within each sample. The datasets met several metrics of high quality and have been deposited to the Gene Expression Omnibus database (GSE202327, GSE202328, GSE202329) as both raw and processed files to serve as long-read reference transcriptomes for future studies of human circulating lymphocytes.

| S-EPMC9635627 | biostudies-literature

Reference Short-Read Transcriptomes of Human Peripheral Blood Lymphocytes (Illumina RNA-Seq, Matched to PacBio Iso-Seq Data)

Project description:Long-read sequencing technologies such as Iso-Seq (PacBio Inc.) generate highly accurate sequences of full-length mRNA transcript isoforms. Long-read transcriptomics may be especially useful in the context of lymphocyte functional plasticity as it relates to human health and disease. However, no long-read isoform-aware reference transcriptomes of human circulating lymphocytes seem to be publicly available despite being valuable as benchmarks in a variety of transcriptomic studies. To begin to fill this gap, we purified four lymphocyte subsets (CD4 T, CD8 T, NK, and Pan B cells) from the peripheral blood of a healthy male donor and obtained high-quality RNA (RIN>8) for PacBio Iso-Seq analysis and parallel RNA-Seq analysis.

2022-09-16 | GSE202328 | GEO

Reference Long-Read Transcriptomes of Human Peripheral Blood Lymphocytes (PacBio Iso-Seq, Matched to Illumina RNA-Seq Data)

2022-09-16 | GSE202327 | GEO

Accurate isoform quantification by joint short- and long-read RNA-sequencing

Project description:This SuperSeries is composed of the SubSeries listed below.

2024-07-10 | GSE271530 | GEO

Reference Short-Read Transcriptomes of Human Peripheral Blood Lymphocytes (Illumina RNA-Seq, Matched to PacBio Iso-Seq Data)

Project description:Reference Short-Read Transcriptomes of Human Peripheral Blood Lymphocytes (Illumina RNA-Seq, Matched to PacBio Iso-Seq Data)

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Targeted DNA-seq and RNA-seq of Reference Samples with Short-read and Long-read Sequencing.

Project description:Next-generation sequencing (NGS) has revolutionized genomic research by enabling high-throughput, cost-effective genome and transcriptome sequencing accelerating personalized medicine for complex diseases, including cancer. Whole genome/transcriptome sequencing (WGS/WTS) provides comprehensive insights, while targeted sequencing is more cost-effective and sensitive. In comparison to short-read sequencing, which still dominates the field due to high speed and cost-effectiveness, long-read sequencing can overcome alignment limitations and better discriminate similar sequences from alternative transcripts or repetitive regions. Hybrid sequencing combines the best strengths of different technologies for a more comprehensive view of genomic/transcriptomic variations. Understanding each technology's strengths and limitations is critical for translating cutting-edge technologies into clinical applications. In this study, we sequenced DNA and RNA libraries of reference samples using various targeted DNA and RNA panels and the whole transcriptome on both short-read and long-read platforms. This study design enables a comprehensive analysis of sequencing technologies, targeting protocols, and library preparation methods. Our expanded profiling landscape establishes a reference point for assessing current sequencing technologies, facilitating informed decision-making in genomic research and precision medicine.

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Reference Long-Read Transcriptomes of Human Peripheral Blood Lymphocytes (PacBio Iso-Seq, Matched to Illumina RNA-Seq Data)

Project description:Reference Long-Read Transcriptomes of Human Peripheral Blood Lymphocytes (PacBio Iso-Seq, Matched to Illumina RNA-Seq Data)

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Reference-free validation of short read data.

Project description:BACKGROUND: High-throughput DNA sequencing techniques offer the ability to rapidly and cheaply sequence material such as whole genomes. However, the short-read data produced by these techniques can be biased or compromised at several stages in the sequencing process; the sources and properties of some of these biases are not always known. Accurate assessment of bias is required for experimental quality control, genome assembly, and interpretation of coverage results. An additional challenge is that, for new genomes or material from an unidentified source, there may be no reference available against which the reads can be checked. RESULTS: We propose analytical methods for identifying biases in a collection of short reads, without recourse to a reference. These, in conjunction with existing approaches, comprise a methodology that can be used to quantify the quality of a set of reads. Our methods involve use of three different measures: analysis of base calls; analysis of k-mers; and analysis of distributions of k-mers. We apply our methodology to wide range of short read data and show that, surprisingly, strong biases appear to be present. These include gross overrepresentation of some poly-base sequences, per-position biases towards some bases, and apparent preferences for some starting positions over others. CONCLUSIONS: The existence of biases in short read data is known, but they appear to be greater and more diverse than identified in previous literature. Statistical analysis of a set of short reads can help identify issues prior to assembly or resequencing, and should help guide chemical or statistical methods for bias rectification.

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LocusMasterTE: long-read assisted short-read TE quantification [short]

Project description:With an ability to compromise genome integrity, transposable elements (TEs) have significant associations with human diseases. Short-read sequencing has been used to study the expression of TEs; however, the highly repetitive nature of these elements makes multimapping a critical issue. Here we implement LocusMasterTE, an improved quantification method by integrating long-read sequencing. Introducing computed transcript per million(TPM) counts from long-read sequencing as prior distribution during Expectation-Maximization(EM) model in short-read TE quantification, multi-mapped reads are re-assigned to correct expression values. Based on simulated short reads, LocusMasterTE outperforms current quantitative approaches and is significantly favorable in capturing newly inserted TEs. We also verified that TEs quantified by LocusMasterTE clearly related to euchromatins and heterochromatins in cell line samples. With LocusMasterTE we anticipate that more accurate quantification can be performed, allowing novel functions of TEs to be uncovered.

2023-09-01 | GSE225380 | GEO

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