Validated DDR1 Lentiviral shRNA #V2261
Knockdown Validated
Aliases
Discoidin Domain Receptor Tyrosine Kinase 1; Epithelial Discoidin Domain-Containing Receptor 1; CD167 Antigen-Like Family Member A; Protein-Tyrosine Kinase RTK-6; Mammary Carcinoma Kinase 10; Tyrosine-Protein Kinase CAK; Cell Adhesion Kinase; Tyrosine Kinase DDR; EC 2.7.10.1; EDDR1; NTRK4; PTK3A; HGK2; RTK6; TRKE; CAK; NEP; Neurotrophic Tyrosine Kinase, Receptor, Type 4; Discoidin Domain Receptor Family, Member 1; Epithelial Discoidin Domain Receptor 1; Discoidin Receptor Tyrosine Kinase; PTK3A Protein Tyrosine Kinase 3A; Neuroepithelial Tyrosine Kinase; Protein-Tyrosine Kinase 3A; CD167a Antigen; EC 2.7.10; MCK-10; CD167; MCK10; TRK E; PTK3; DDR
Background
NCBI Gene Entry: 780
Application Information
Human DDR1 mRNA knockdown
Storage Buffer
Culture medium
Storage Instruction
Product should be stored at –80°C
Recommended Dilution
1:2
Protocols
Read all instructions carefully before using this product
Instructions For Use
Note: The following protocol uses HeLa cell as an example assuming that your cell culture medium is DMEM-based. If you use a different cell culture medium, please contact us for a special quote or technical support.
- Release 0.5 million HeLa cells into a 35 mm tissue culture dish in 2 mL of the growth medium (DMEM containing 10% FBS and 1% pen/strep). Cell density should reach 50-60% confluence the following day.
- 24 h after cell release, pre-warm the shRNA lentiviral medium to 37°C.
- Discard 1 mL of the original growth medium of the 35 mm dish.
- Using a serological pipette, gently mix the lentiviral solution 3 times.
- Carefully add 1 mL of the lentiviral solution to the well. Tip: To prevent splashing, add the solution to the dish along the wall.
- Add a polybrene stock solution to the culture medium at a final concentration of 5 μg/mL. Gently swirl the dish to mix.
- 48 h after cell release, without discarding the original medium, add another 1mL of lentiviral medium directly into the dish.
- Add an additional polybrene stock solution into the dish to obtain a final concentration of 5 μg/mL. Tip: Now, the medium in the dish should be a total of 3 mL.
- 72 h after cell release, cells may reach confluence. Trypsinize the cells off the 35 mm dish and culture those cells in a 60 mm dish.
- Add puromycin to the dish at a final concentration of 4 μg/mL. Tip: To assess the efficacy of puromycin selection, culture a dish of wild-type HeLa cells as a negative control.
- Allow puromycin selection for 48 h. Almost all wild-type HeLa cells should die, while the dish infected with lentiviruses should have some remaining cells.
- Replace the medium with the regular growth medium without puromycin and allow the cells to grow to confluence before harvesting or staining.