|
| Status |
Public on Feb 09, 2023 |
| Title |
Plasmid replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
plasmid DNA
|
| Organism |
synthetic construct |
| Characteristics |
tissue: plasmid DNA
|
| Treatment protocol |
K562 cells 10 million cells per replicate were transfected with 40ug the MPRA library using electroporation with the Neon transfection system following the manufacturers protocol.
|
| Growth protocol |
K562 cells were grown with RPMI 1640 Complete Medium is supplemented with 2mM L-Glutamine + 10% FBS + 1% Penicillin-Streptomycin to a density of ~6*10^5 cells/ML
|
| Extracted molecule |
other |
| Extraction protocol |
Twenty-four hours after transfection, cells were collected and snap frozen in liquid N2. Total RNA was extracted and GFP mRNA was isolatedand converted to cDNA Tewhey and colleagues 2016.
Following Tewhey et al. 2016, cDNA libraries were quanitified using qPCR including the plasmid pool used to transfect the cells. They were then diluted to be of equal concentration with the lowest library (20 cycles) and amplified using one fewer cycle than the lowest cycle threshold. An additional PCR was performed to add P7 indexing primers.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina MiSeq |
| |
|
| Description |
MPRA Plasmid DNA
covid_mpra_count_file_forDeseq.txt
|
| Data processing |
Library strategy: MPRA
Barcodes were extracted from sequence files and matched to oligos using the attached covid_mpra_plasmid_seq_outfile_ALL_4_INDEXES.txt.gz file. The barcode is the first column and the oligos attached to it are in the last column. Any barcodes matched to duplicate oligos were excluded. Oligo sequences that match the oligo ids can be found in the attached covid_oligo_pool_just_oligos_and_names_10.14.20.txt (column 1 is the oligo sequence, column 2 is the oligo id).
Counts per oligo per replicate were determined by summing all barcode matches together for each replicate and a reported unnormalized in the included count file.
Supplementary_files_format_and_content: Counts per oligo per replicate were determined by summing all barcode matches together for each replicate and are reported unnormalized in the included count file.
|
| |
|
| Submission date |
Jun 06, 2021 |
| Last update date |
Feb 09, 2023 |
| Contact name |
Terence D Capellini |
| E-mail(s) |
[email protected], [email protected]
|
| Phone |
6173010673
|
| Organization name |
Harvard Univserity
|
| Department |
Human Evolutionary Biology
|
| Lab |
Capellini
|
| Street address |
11 Divinity Avenue
|
| City |
Cambridge |
| State/province |
MA |
| ZIP/Postal code |
02138 |
| Country |
USA |
| |
|
| Platform ID |
GPL17769 |
| Series (1) |
| GSE176233 |
Regulatory dissection of the severe COVID-19 risk locus introgressed by Neanderthals. |
|
| Relations |
| BioSample |
SAMN19588370 |
| SRA |
SRX11078208 |
