The scRNA-seq datas of human umbilical cord mesenchymal stromal cells (22+5 W and 28 W)

Published: 6 July 2023| Version 1 | DOI: 10.17632/36tfjc42hm.1
Contributor:
Chuiqin Fan

Description

Cell Preparation: HUMSCs in passage 3 from three different gestational ages (22+5 W, 28 W) were used in this study. Cell survival rates were checked and single-cell suspensions with survival rates above 80% were washed and resuspended to a suitable concentration of 700-1200 cells/μl for 10x Genomics Chromium (10X Genome, Single Cell 3’ Library & Gel Bead Kit v.3). The system was operated on the machine. GEM Creation & Thermal Cycling: Gel Bead in Emulsion (GEMs) were created for single-cell separation according to the number of cells to be harvested. Once the GEMs were formed, they were collected for reverse transcription in a PCR machine for labeling.Post Cycling Cleanup & cDNA Amplification: The GEMs were treated with oil and the amplified cDNA was purified by magnetic beads. It then underwent cDNA amplification and quality inspection. Library Preparation & Quantification: The 3ʹ Gene Expression Library was constructed with the quality-qualified cDNA. After fragmentation, adapter ligation, sample index PCR, etc., the library was finally quantitatively examined.Sequencing: The final library pool was sequenced on the Illumina HiSeq3000 using 150-base-pair paired-end reads.

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Steps to reproduce

Cell Preparation: HUMSCs in passage 3 from three different gestational ages (22+5 W, 28 W) were used in this study. Cell survival rates were checked and single-cell suspensions with survival rates above 80% were washed and resuspended to a suitable concentration of 700-1200 cells/μl for 10x Genomics Chromium (10X Genome, Single Cell 3’ Library & Gel Bead Kit v.3). The system was operated on the machine. GEM Creation & Thermal Cycling: Gel Bead in Emulsion (GEMs) were created for single-cell separation according to the number of cells to be harvested. Once the GEMs were formed, they were collected for reverse transcription in a PCR machine for labeling.Post Cycling Cleanup & cDNA Amplification: The GEMs were treated with oil and the amplified cDNA was purified by magnetic beads. It then underwent cDNA amplification and quality inspection. Library Preparation & Quantification: The 3ʹ Gene Expression Library was constructed with the quality-qualified cDNA. After fragmentation, adapter ligation, sample index PCR, etc., the library was finally quantitatively examined.Sequencing: The final library pool was sequenced on the Illumina HiSeq3000 using 150-base-pair paired-end reads.

Institutions

China Medical University

Categories

Single-Cell RNA Sequencing

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