siRNA / miRNA gene silencing Human - MDA-MB-231 LAT1/SLC7A5

miRNA is the inherent gene silencing machinery which can have more than one mRNA target, whereas siRNA can be designed to target a particular mRNA target. By design, both siRNA and miRNA are 20-25 nucleotides in length. The target sequence for siRNAs is usually located within the open reading frame, between 50 and 100 nucleotides downstream of the start codon. There are two ways in which cells can be transfected with desired RNAi: 1. Direct transfection (with calcium phosphate co-precipitation or cationic lipid mediated transfection using lipofectamine or oligofectamine), and 2. Making RNAi lentiviral constructs (followed by transformation and transduction). Lentiviral constructs are time consuming, but provide a more permanent expression of RNAi in the cells, and consistent gene silencing. Direct transfection of oligonucleotides provides temporary genetic suppression. Traditional methods like calcium phosphate co-precipitation have challenges like low efficiency, poor reproducibility and cell toxicity. Whereas, cationic lipid-based transfection reagents are able to overcome these challenges, along with applicability to a large variety of eukaryotic cell lines. When using oligos, the ideal concentration lies between 10-50nM for effective transfection.

1 Matching solution
Start a discussion

Start discussion

No discussions found

Start your discussion

Share your thoughts or question with experts in your field

Start a discussion

Found 1 matching solution for this experiment

ON-TARGETplus Human SLC7A5 (8140) siRNA - Individual

Horizon Discovery Ltd.

Upstream tips	Protocol tips	Downstream tips
	Non-targeting short interfering RNA (siRNA) (cat # D-001810-01-20), ON-TARGET plus human siRNA's against AHR (cat # J-004990-08-0010, and cat # J-004990-06-0010) and LAT1 (cat # J-004953-09-0010) were purchased from GE Dharmacon (Lafayette, CO). MDA-MB-231 BCCs were purchased from ATCC (Manassas, VA) and maintained in DMEM, 10% FBS, with penicillin (100 IU/mL) and streptomycin (100 IU/mL). The siRNA knockdowns were performed as detailed in our prior reports [33, 34]. Briefly, 200,000 cells (MDA-MB-231) in 1 mL of DMEM supplemented with 10% FBS were mixed directly with 100 nM of siRNA that was non-targeting, AHR-targeting or LAT1-targeting and 3 μL of Lipofectamine RNAiMAX transfection reagent (Thermo Fisher Scientific) and immediately plated in 35 mm tissue culture plates for 48 hrs.

Protocol tips
Non-targeting short interfering RNA (siRNA) (cat # D-001810-01-20), ON-TARGET plus human siRNA's against AHR (cat # J-004990-08-0010, and cat # J-004990-06-0010) and LAT1 (cat # J-004953-09-0010) were purchased from GE Dharmacon (Lafayette, CO). MDA-MB-231 BCCs were purchased from ATCC (Manassas, VA) and maintained in DMEM, 10% FBS, with penicillin (100 IU/mL) and streptomycin (100 IU/mL). The siRNA knockdowns were performed as detailed in our prior reports [33, 34]. Briefly, 200,000 cells (MDA-MB-231) in 1 mL of DMEM supplemented with 10% FBS were mixed directly with 100 nM of siRNA that was non-targeting, AHR-targeting or LAT1-targeting and 3 μL of Lipofectamine RNAiMAX transfection reagent (Thermo Fisher Scientific) and immediately plated in 35 mm tissue culture plates for 48 hrs.

Protocol tips

Non-targeting short interfering RNA (siRNA) (cat # D-001810-01-20), ON-TARGET plus human siRNA's against AHR (cat # J-004990-08-0010, and cat # J-004990-06-0010) and LAT1 (cat # J-004953-09-0010) were purchased from GE Dharmacon (Lafayette, CO). MDA-MB-231 BCCs were purchased from ATCC (Manassas, VA) and maintained in DMEM, 10% FBS, with penicillin (100 IU/mL) and streptomycin (100 IU/mL). The siRNA knockdowns were performed as detailed in our prior reports [33, 34]. Briefly, 200,000 cells (MDA-MB-231) in 1 mL of DMEM supplemented with 10% FBS were mixed directly with 100 nM of siRNA that was non-targeting, AHR-targeting or LAT1-targeting and 3 μL of Lipofectamine RNAiMAX transfection reagent (Thermo Fisher Scientific) and immediately plated in 35 mm tissue culture plates for 48 hrs.

Manufacturer protocol Publication protocol 1 Relevant paper

Can't find the product you've used to perform this experiment? It would be great if you can help us by Adding a product!

Outsource your experiment

Fill out your contact details and receive price quotes in your Inbox

Outsource experiment