|
| Status |
Public on Apr 20, 2019 |
| Title |
alien2_dna: fully_randomized_2_alien2 plasmid DNA sequencing |
| Sample type |
SRA |
| |
|
| Source name |
plasmid alternative polyadenylation reporter
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: HEK293T
passages: 10
molecule: reporter DNA
|
| Treatment protocol |
Plated cells were seeded on ECM-coated plates and grown to ~90% confluency for 24-36 hours. Plasmid DNA was complexed with Lipofectamine 3000 reagent and transiently transfected.
|
| Growth protocol |
HEK293T cells were cultured on ECM-coated plates in Dubelco's DMEM media supplemented with FBS (5%), penicillin (.05%), and streptomycin (.05%)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from transiently transfected cells 36-48 hours later using the Qiagen Rneasy kits. The purified RNA was then polyA purified using the NEB PolyA Spin kit (magnetic-bead-based protocol).
Library specific priming sites were used to append Illumina adapter sequences and to amplify the library. Amplification was conducted in a light-cycler and samples removed early to minimize PCR bias.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 550 |
| |
|
| Data processing |
DNA and RNA sequences distinguished using internal sequence tags
Sequences sorted based on human-derived background sequence tags
DNA and full length (long isoform) RNA used to build sequence map
RNA mapped back to the full-length sequences and the isoform and site of polyadenylation recorded
RNA short and long isoform ratios calculated for each unique library sequence
Genome_build: Reporter UTR background sequence designs based on Human build 37 GRCh37
Supplementary_files_format_and_content: CSV files contain reporter 3'UTR sequences and isoform calculations. Numpy matrix files contain polyadenylation site positions.
|
| |
|
| Submission date |
Apr 30, 2018 |
| Last update date |
Apr 20, 2019 |
| Contact name |
Georg Seelig |
| E-mail(s) |
gseelig@uw.edu
|
| Organization name |
University of Washington
|
| Department |
Electrical Engineering
|
| Street address |
3946 W Stevens Way NE
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98195-0001 |
| Country |
USA |
| |
|
| Platform ID |
GPL21697 |
| Series (1) |
| GSE113849 |
Degenerate 3'UTR DNA and RNA sequence data |
|
| Relations |
| BioSample |
SAMN09001089 |
| SRA |
SRX4012745 |
