|
| Status |
Public on May 01, 2024 |
| Title |
Parasrampuria_mRNA_Seq_shHD-33full_UAC_1 |
| Sample type |
SRA |
| |
|
| Source name |
Coriell Institute HD fibroblasts (GM04281)
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Coriell Institute HD fibroblasts (GM04281)
cell type: Fibroblast
biomaterial provider: https://catalog.coriell.org/0/Sections/Search/Sample_Detail.aspx?Ref=GM04281&product=CC
genotype: HTT-Patient
treatment: Infection with lentiviral particles encoding smRNA
|
| Treatment protocol |
Cells were infected with lentivirus vectors encoding smRNA at a MOI of 5 containing 1 ug/mL polybrene and incubated at 37°C in a humidified 5% CO incubator, changing media regularly.
|
| Growth protocol |
HD fibroblasts were cultured in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum (Gibco), 1x non-essential amino acids (100X, Gibco), and 1% sodium pyruvate (100X, Gibco) at 37°C in 5% CO2.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was Trizol extracted 7 days following infection and poly-A was isolated using magnetic bead purification methods.
The isolated mRNA was prepared using the cDNA-PCR Barcoding Kit V14 (Oxford Nanopore Technologies) following the manufacturers protocol and sequenced on a PromethION 2 Solo instrument (ONT).
RNA-Seq by cDNA-PCR, short read sequencing by Oxford Nanopore P2 Solo methods
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
PromethION |
| |
|
| Description |
Oxford Nanopore Technologies Promethion 2 Solo
Parasrampuria_DESeq2_gene_counts.csv
Parasrampuria_Top_Gene_Change_Summary.csv
Parasrampuria_DESeq2_results_shHD33fullUAC.csv
|
| Data processing |
Sequenced reads were basecalled and demultiplexed using Dorado (v0.5.1).
Minimap2 alignment software was used to align demultiplexed reads to the GENCODE transcriptome reference Release 45 (GRCh38.p14).
Bam files were quantified using the Salmon RNA transcript quantification.
RNA-Seq data was analyzed using the DESeq2 (v1.42.1) package following the transcript abundance and tximport/tximeta input data format.
DESeq2 analysis codes are available in the supplementary file section.
GENCODE transcriptome reference Release 45 (GRCh38.p14)
Assembly: Parasrampuria_DESeq2_gene_counts.csv: Raw gene counts found using DESeq2 between sequenced samples using genes of interest.
Supplementary files format and content: Parasrampuria_Top_Gene_Change_Summary.csv: Top ten genes differentially expressed determined by greatest p-adj value normlaized to shMCHR control.
Supplementary files format and content: Parasrampuria_DESeq2_results_XXX.csv: DESeq2 results output to .csv with GeneIDs. Gene differentiation normalized to shMCHR control.
|
| |
|
| Submission date |
Apr 17, 2024 |
| Last update date |
May 01, 2024 |
| Contact name |
Keith T. Gagnon |
| E-mail(s) |
ktgagnon@wakehealth.edu
|
| Organization name |
Wake Forest University School of Medicine
|
| Department |
Biochemistry and Molecular Biology
|
| Street address |
575 Patterson Ave
|
| City |
Winston-Salem |
| State/province |
North Carolina |
| ZIP/Postal code |
27101 |
| Country |
USA |
| |
|
| Platform ID |
GPL26167 |
| Series (2) |
| GSE264218 |
Sequencing-Guided Design of Genetically Encoded Small RNAs Targeting CAG Repeats for Selective Inhibition of Mutant Huntingtin [II] |
| GSE264444 |
Sequencing-Guided Design of Genetically Encoded Small RNAs Targeting CAG Repeats for Selective Inhibition of Mutant Huntingtin |
|
| Relations |
| BioSample |
SAMN40995347 |
| SRA |
SRX24288784 |
