Table 2.
A list of target sequences for sgRNAs and shRNA.
| Antibody | Catalog Number | Source |
|---|---|---|
| Anti-BCKDK (E-12) | sc-374425 | Santa Cruz Technology (CA, USA) |
| Anti-Fyn | sc-434 | |
| Anti-MEK1/2 | #8727 | Cell Signaling Technology (Danvers, MA, USA) |
| Anti-p-MEK1/2 (S221) | #2338 | |
| Anti-His | #2365s | |
| Anti-HA-tag | #3724 | |
| Anti-p-BCKDHA (S293) | ab200577 | Abcam (Cambridge, UK) |
| Anti-BCKDHA | ab68094 | |
| Anti-BCKDK | PA5-86367 | Thermo Fisher (Waltham, MA, USA) |
| Anti-Alpha Tubulin | 66031-1-Ig | Proteintech (Wuhan, China) |
| Anti-Myc-tag | 16286-1-AP | |
| Anti-Myc-tag | 60003-2-Ig | |
| Anti-GST-tag | 66001-2-Ig | |
| Anti-ubiquitin | 10201-2-AP | |
| Anti-ACY1 | 11316-1-AP | Proteintech (Wuhan, China) |
| Anti-p-Thr | AB1607 | Sigma-Aldrich (St. Louis, MO, USA) |
| Anti-Ser | 05-1000 | |
| Anti-Flag | F7425 | |
| Anti-Flag | F1804 | |
| Anti-p-BCKDK (Y151) | Abiocode (Shanghai, China) | |
| HRP-conjugated secondary antibodies (R) | E030120-2 | EarthOx Life (San Francisco, CA, USA) |
| HRP-conjugated secondary antibodies (M) | E030110-02 |
| Regent | Catalog Number | Source |
|---|---|---|
| 3,6-dichlorobenzothiophene-2-carboxylic acid (BT2) | ZC-26488 | ZZBIO.CO.LTD (Shanghai, China) |
| γ-32P-ATP | BLU002H250UC | Perkin Elmer (Beijing, China) |
| inactive human BCKDK recombinant protein | (TP303601) | Beijing OriGene Technology |
| Fyn active kinase | #14-441 | Millipore Corp (Billerica, MA, USA) |
| Cycloheximide (CHX) | C7698-5G | Sigma-Aldrich (St. Louis, MO, USA) |
| Polybrene | TR-1003-G | |
| Puromycin | B7587 | APExBio (Houston, USA) |
| G418 was purchased from GLPBIO (Montclair, CA) | GC17427 | GLPBIO (Montclair, CA) |
| N-acetyl-L-alanine | S20262 | Shyuanye (Shanghai, China) |
| D-Luciferin sodium salt | S19261 | |
| Isopropyl β-D-thiogalactoside (IPTG) | ST098 | Beyotime (Shanghai, China) |
| Plasmid | Catalog Number | Source |
|---|---|---|
| pcFlag-Fyn-WT | #74509 | addgene (Cambridge, MA, USA) |
| phage | #118692 | |
| lentiCRISPR V2 | #52961 | |
| pCMV-BCKDK-Flag WT | Laboratory owned and self-constructed | |
| pCMV-BCKDK-Flag Y151F | ||
| pcDNA4-His-BCKDK | ||
| pcDNA4-His-BCKDK Y151F | ||
| phage-Myc-BCKDK-Flag WT | ||
| phage-Myc-BCKDK-Flag Y151F | ||
| phage-Myc-BCKDK-Flag Y151D | ||
| pET23a-BCKDK-His |