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Single-Cell RNA Sequencing

Service Overview
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single-cell-rna-sequencingSingle-cell sequencing is now readily available to researchers through Novogene’s expertise in this field. Novogene is one of the few NGS providers with extensive experience in single-cell sequencing technology, including single-cell RNA-Seq. We offer the highest quality services in amplification, library construction, sequencing, and bioinformatics analysis to our customers, and our results have been published in leading scientific journals.

Single-cell RNA-Seq enables the high-resolution transcriptome profiling of a single cell, and has broad utility for investigating developmental processes and gene regulatory networks, and for revealing heterogeneous gene expression patterns within cell cultures, tissues, and organs. Applications include profiling gene expression changes during stem cell differentiation, organ development, or tumor progression, and characterizing the response of subpopulations of cells to environmental signals and conditions.

Novogene offers the most advanced techniques for transcriptome analyses for single cells.

The Novogene Advantage

  • Leader in single-cell RNA-Seq: We are one of the few providers of this technology, with the highest ranking in technical capability and experience, and publications in the field.
  • Advanced amplification methods: We use the SMARTer kit for single cell RNA amplification, which reduces amplification costs, improves amplification rates, and has been utilized in multiple publications. The success rate for amplification from human peripheral blood lymphocytes and mammalian embryonic cells are 100% and >62%, respectively.
  • Comprehensive processing: Our single-cell RNA-Seq service includes amplification, library construction, sequencing, and bioinformatics analysis.

Project Workflow

single cell rna seq service project workflow

Sequencing Strategy

  • 250-300 bp insert cDNA library
  • HiSeq platform, paired-end 150 bp

Data Quality Guarantee

  • Q30 ≥ 80%

Sample Requirements

  • Sorted single cells should be stored in one of the two ways (1) in lysis buffer with RNase inhibitor from SMARTer kit (Clontech) or (2) in 1X PBS buffer (excluding calcium and magnesium) containing RNase inhibitor in a total volume of ≤ 2 μl. The stored cell should be frozen in liquid nitrogen and shipped out with dry ice.

Turnaround Time

  • Amplification: 7 working days from verification of sample quality
  • Library preparation and sequencing: within 15 working days
  • The turnaround for data analysis is project dependent

Analysis Pipeline

single-cell rna seq service analysis pipeline

Table. Representative data quality results of single-cell RNA-Seq service from Novogene.

Sample Raw reads Clean reads Clean bases Error (%) Q 20 (%) Q 30(%) GC (%)
sample1_1 20594409 20132577 2.52G 0.03 96.16 92.32 43.60
sample1_2 20594409 20132577 2.52G 0.04 93.57 88.31 43.56
sample2_1 22213754 21739346 2.72G 0.03 96.22 92.38 43.69
sample2_2 22213754 21739346 2.72G 0.04 93.90 88.87 43.65

Project Example

The following studies utilized Novogene’s expertise in single-cell RNA-Seq.

Tracing haematopoietic stem cell formation at single-cell resolution
Nature 533: 487-492 (2016)

In this study, Novogene’s advanced single-cell RNA sequencing technology was used to survey the transcriptomes of pre-HSCs (haematopoietic stem cells) and HSCs during embryonic development at single-cell and single-base resolution. Specifically, the researchers discovered that mTOR (mechanistic targets of rapamycin) signaling is indispensable for the emergence of HSCs but not haematopoietic progenitors. The pre-HSCs showed remarkable heterogeneity in cell-cycle status. Overall, the combination of single-cell RNA sequencing with single-cell capture markers successfully provided key information for engineering HSCs for clinical applications.

single cell rna fig 1
Figure. Global gene expression dynamics during HSC formation

Single-cell RNA-Seq analysis maps development of human germline cells and gonadal niche interactions
Cell Stem Cell 20(6): 858-873 (2017)

In this study, Novogene’s advanced single-cell RNA-Seq service was used on 2,167 single fetal germ cells (FGCs) from male and female human embryos to explore the developmental stages and heterogeneity of FGCs. Researchers identified four developmental stages for female FGCs and three for male FGCs. Reciprocal signaling in the NOTCH and BMP pathways was also found to occur between the FGCs and their gonadal niche cells. Furthermore, key candidate master transcription factors were identified, which are crucial for stage-specific development in both male and female FGCs. This study, as the first to perform such a comprehensive analysis on human FGCs, demonstrates the powerful capabilities of single-cell RNA sequencing and its unlimited potential in further exploring human germ cell development.

Figure. Dynamic Gene Expression Patterns of Human Female FGCs

Single-cell RNA-Seq profiling of human preimplantation embryos and embryonic stem cells.
Nature Structural & Molecular Biology, 20:1131-1139 (2013).

In this study, the transcription profiles of human embryonic stem cells and cells from human preimplantation embryos were examined at different developmental stages. Results demonstrated markedly different expression patterns between cell groups, and revealed over 2,000 novel long noncoding RNAs.

Figure. Expression patterns during human preimplantation development and derivation of human embryonic stem cells (hESCs)

Figure. Number and spectrum of SNVs

Examples of Publications Using Novogene’s Services

Journal Title
Nature Structural & Molecular Biology, 20: 1131-1139 (2013) Single-cell RNA-Seq profiling of human preimplantation embryos and embryonic stem cells.
Nature, 533: 487-492 (2016) Tracing haematopoietic stem cell formation at single-cell resolution.
Cell Stem Cell, 20(6): 858-873 (2017) Single-cell RNA-Seq analysis maps development of human germline cells and gonadal niche interactions.
Cell Stem Cell, 21(5): 635-649.e8 (2017) The primate-specific gene TMEM14B marks outer radial glia cells and promotes cortical expansion and folding.
Cell Research, 28: 730–745 (2018) Spatial transcriptomic survey of human embryonic cerebral cortex by single-cell RNA-seq analysis.
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