|
| Status |
Public on Jul 12, 2024 |
| Title |
DHD0 (subset), DHD2, mALb (T) His+ |
| Sample type |
SRA |
| |
|
| Source name |
yeast cell
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
cell type: yeast cell
genotype: MATa AGA1::GAL1-AGA1::URA3 ura3-52 trp1 leu2-delta200 his3-delta pep4::BleoMX leu2::pGAL6z-HIS3 prbd1.6R can1 GAL
|
| Growth protocol |
250uL of transformed yeast from gylcerol stock is first grown in 100mL of synthetic drop out media at 30C and 250rpm lacking tryptophan during an extended outgrowth step. Yeast are allowed to grow for at least 36 hours with periodic 1:250 dillutions into fresh media when optical density at 600nM reaches 1.0. 250uL of cells are used to inoculate 100mL of synthetic dropout media - Tryptophan -Histidine +3mM 3AT. Cells are allowed to incubate at same conditions before until OD600 of 1.0 is reached.
|
| Extracted molecule |
other |
| Extraction protocol |
All plasmids are extracted after growth selection using a Zymoprep Yeast Plasmid Miniprep II Kit according to the manufacturer's protocol.
All proteins of interest are ordered as twist gene fragements. These fragments are ordered to contain the terminator, barcode, and overlap regions for assembly. These fragmetns are combined with the MP3-seq vector and assembled during yeast transformation through homologous recombination.
|
| |
|
| Library strategy |
OTHER |
| Library source |
other |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
| |
|
| Description |
An all-by-all PPI assay collected with version one of MP3-Seq. 1mM 3-AT.
L43 His+
|
| Data processing |
Trimmed fastq files are clustered with starcode
Clustered barcodes are matched to their proteins using the barcode to protein tables (see supplemental file), and PPIs are counted.
Autoactivators are detected and corrected (see methods of paper)
Deseq2 is used to get fold changes for each PPI using the His+ and His- condition counts as inputs.
Assembly: N/A
Supplementary files format and content: Proccessed PPI counts per each selection stage (His+ and His-)
The first column of the csv file has the DBD associated protein name, while the columns have the AD associated protein name. The values in all other cells correspond to the count of the barcodes for the PPI being measured.
Library strategy: MP3-seq
|
| |
|
| Submission date |
Jul 08, 2024 |
| Last update date |
Jul 12, 2024 |
| Contact name |
Alyssa La Fleur |
| Organization name |
University of Washington
|
| Lab |
Seelig Lab
|
| Street address |
Campus Box 352500 185 Stevens Way
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98195-2500 |
| Country |
USA |
| |
|
| Platform ID |
GPL26302 |
| Series (1) |
| GSE271790 |
MP3-seq: Massively parallel measurement of protein-protein interactions by sequencing |
|
| Relations |
| BioSample |
SAMN42390919 |
| SRA |
SRX25261434 |
