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Status |
Public on Jan 31, 2017 |
Title |
Drop-Seq analysis of Pooled Arc-ME single-cell suspension_ad lib 60% high fat diet fed_1 |
Sample type |
SRA |
|
|
Source name |
Arcuate Nucleus/Median Eminence
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Organism |
Mus musculus |
Characteristics |
strain: C57BL6/J tissue: Arcuate-Median Eminence Sex: M average age: 47 number of mice pooled: 4 experimental condition: Ad lib fed 60% High fat diet batch: Batch 4
|
Extracted molecule |
total RNA |
Extraction protocol |
Single-cell suspensions were processed through Drop-Seq to generate single-cell cDNA libraries attached to microbeads. Microbeads were counted, and amplified by PCR, and the 3' end of the cDNA prepared for sequencing using a modified Nextera XT protocol.
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NextSeq 500 |
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Description |
HFD Cell Barcodes are in read 1, gene sequence in read 2,
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Data processing |
Base calls performed with bcl2fastq2.15.0.4 Read 2 was aligned with Star v2.4.0. Each aligned read was tagged with its mate-pair UMI and cell barcode. Each aligned exonic read was tagged with a gene name. A digital expression matrix was built by counting the number of unique UMIs per gene within each cell; columns are cells, and rows are genes. The first inflection point was estimated on a cumulative sum plot of the digital expression matrix to determine the number of cells in the sample. Cells expressing >800 genes from Samples 1-11 were merged together into a single matrix. Merged DGE file from all 11 samples generated. log(1+10,000*(DGE normalized to total transcripts per cell)) calculated on DGE. This is the Merged_all_020816_DGE.txt.gz file containing 21,421 cell barcodes Genes showing expression in <50 cells were removed and batch effect correction was performed using the removeBatchEffect function of edgeR Data was Natural log transformed and cells were clustered using Seurat software (PCA followed by t-SNE and density clustering) Cell clusters containing markers for more than one other cluster were considered doublets and were removed. Remaining cells are presented in manuscript and contained in Merged_all_020816_BatchCorrected_LNtransformed_doubletsremoved_Data.txt file Genome_build: Genome_build: mm10 (GRCm38) Supplementary_files_format_and_content: Digital expression matrices. Compressed, flat text files with tab delimiters. Supplementary_files_format_and_content: Merged_all_020816_DGE.txt: Digital Gene Expression Matrix of raw data Supplementary_files_format_and_content: Merged_all_020816_BatchCorrected_LNtransformed_doubletsremoved_Data.txt: Digital Gene Expression Matrix, Batch corrected,natural log transformed, cell doublets removed Supplementary_files_format_and_content: cell_metadata.txt: Metadata for individual cells based on final cluster assignments
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Submission date |
Jan 10, 2017 |
Last update date |
May 15, 2019 |
Contact name |
Linus Tzu-Yen Tsai |
E-mail(s) |
ltsai@bidmc.harvard.edu
|
Organization name |
Beth Israel Deaconess Medical Center
|
Department |
Medicine, Division of Endocrinology, Diabetes, and Metabolism
|
Street address |
330 Brookline Avenue
|
City |
Boston |
State/province |
MA |
ZIP/Postal code |
02115 |
Country |
USA |
|
|
Platform ID |
GPL19057 |
Series (1) |
GSE93374 |
A Molecular Census of Arcuate Hypothalamus and Median Eminence Cell Types |
|
Relations |
BioSample |
SAMN06215611 |
SRA |
SRX2481512 |