SNAI 1 siRNA and shRNA Plasmids (h)

siRNA / miRNA gene silencing Human - MDA-MB-231 SNAI 1

Experiment
siRNA / miRNA gene silencing Human - MDA-MB-231 SNAI 1
Product
SNAI 1 siRNA and shRNA Plasmids (h) from Santa Cruz Biotechnology
Manufacturer
Santa Cruz Biotechnology

Protocol tips

Protocol tips
MDA-MB-468 breast adenocarcinoma cell lines were purchased from the Pasteur Institute, Tehran, Iran, and was maintained in RPMI-1640 medium supplemented with 10% FBS, 1% antibiotics (100 unit/ml penicillin and 100 μg/ml streptomycin). Cell lines were cultured at 37 °C in a humidified atmosphere containing 5% carbon dioxide. Every day we changed the culture medium and passaged the cells when they reached 80% to 90% confluences. Snail1 targeting siRNAs were designed by using Santacruz Biotechnology California, USA (http://www.scbt.com/datasheet-38398-snai-1-sirna-h.html). This designed siRNA sequences were blasted against the human genome database to eliminate cross-silence phenomenon with non-target genes. Scrambled siRNA (Santacruz) that does not target any gene was used as the negative control siRNA. Pooled human snail1 siRNA is a pool of 3 different siRNA duplexes sequences including siRNA duplex A, Sense: GGACUUUGAUGAAGACCAUtt and Antisense: AUGGUCUUCAUCAAAGUCCtt, siRNA duplex B, Sense: CACGAGGUGUGACUAACUAtt and Antisense: UAGUUAGUCACACCUCGUGtt, siRNA duplex C, Sense: GCGAGCUGCAGGACUCUAAtt and Antisense: UUAGAGUCCUGCAGCUCGCtt. We needed to acquire the time and dosage affected by the siRNA. To do it average dosage of 60 was treated in three times of 24, 48 and 72 h to gain the effective time and three dosages of 40, 60 and 80 pmol were evaluated in the gained effective time to acquire the effective dosage. Cells were transfected with siRNA and transfection reagent according to the manufacturer’s instructions. Transfection of siRNA was done in three doses of 40, 60 and 80 pmol. Briefly, cells were seeded in a 6-well-plate at a density of 1 × 106 cells/well with antibiotics-free medium 40 min before the transfection. Six μl of the siRNA concentration was mixed with 6 μl transfection reagent in 200 μl optimal medium and were incubated at room temperature for 30 min to form a complex. After rinsing cells with PBS, 212 μl transfection mixtures were supplemented to each well with 800 μl optimal medium. Six hours after the transfection, the medium was replaced with fresh 1 ml RPMI-1640 medium containing 20% FBS. Overall, 24, 48 and 72 h after the transfection, cells was collected for RNA and protein isolation.

Publication protocol

MDA-MB-468 breast adenocarcinoma cell lines were purchased from the Pasteur Institute, Tehran, Iran, and was maintained in RPMI-1640 medium supplemented with 10% FBS, 1% antibiotics (100 unit/ml penicillin and 100 μg/ml streptomycin). Cell lines were cultured at 37 °C in a humidified atmosphere containing 5% carbon dioxide. Every day we changed the culture medium and passaged the cells when they reached 80% to 90% confluences. Snail1 targeting siRNAs were designed by using Santacruz Biotechnology California, USA (http://www.scbt.com/datasheet-38398-snai-1-sirna-h.html). This designed siRNA sequences were blasted against the human genome database to eliminate cross-silence phenomenon with non-target genes. Scrambled siRNA (Santacruz) that does not target any gene was used as the negative control siRNA. Pooled human snail1 siRNA is a pool of 3 different siRNA duplexes sequences including siRNA duplex A, Sense: GGACUUUGAUGAAGACCAUtt and Antisense: AUGGUCUUCAUCAAAGUCCtt, siRNA duplex B, Sense: CACGAGGUGUGACUAACUAtt and Antisense: UAGUUAGUCACACCUCGUGtt, siRNA duplex C, Sense: GCGAGCUGCAGGACUCUAAtt and Antisense: UUAGAGUCCUGCAGCUCGCtt. We needed to acquire the time and dosage affected by the siRNA. To do it average dosage of 60 was treated in three times of 24, 48 and 72 h to gain the effective time and three dosages of 40, 60 and 80 pmol were evaluated in the gained effective time to acquire the effective dosage. Cells were transfected with siRNA and transfection reagent according to the manufacturer’s instructions. Transfection of siRNA was done in three doses of 40, 60 and 80 pmol. Briefly, cells were seeded in a 6-well-plate at a density of 1 × 106 cells/well with antibiotics-free medium 40 min before the transfection. Six μl of the siRNA concentration was mixed with 6 μl transfection reagent in 200 μl optimal medium and were incubated at room temperature for 30 min to form a complex. After rinsing cells with PBS, 212 μl transfection mixtures were supplemented to each well with 800 μl optimal medium. Six hours after the transfection, the medium was replaced with fresh 1 ml RPMI-1640 medium containing 20% FBS. Overall, 24, 48 and 72 h after the transfection, cells was collected for RNA and protein isolation.

Full paper   Login or join for free to view the full paper.

Reviews

SNAI 1 siRNA and shRNA Plasmids (h) from Santa Cruz Biotechnology has not yet been reviewed for this experiment

We'd love it if you would be the first to write a review!

Discussion

Start your discussion

Share your thoughts or question with experts in your field

Start a discussion

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing siRNA / miRNA gene silencing Human - MDA-MB-231 SNAI 1 using SNAI 1 siRNA and shRNA Plasmids (h) from Santa Cruz Biotechnology.

View full paper   Login or join for free to view the full paper.

Manufacturer protocol

Download the product protocol from Santa Cruz Biotechnology for SNAI 1 siRNA and shRNA Plasmids (h) below.

Download PDF Download manufacturer protocol

Videos

Check out videos that might be relevant for performing siRNA / miRNA gene silencing Human - MDA-MB-231 SNAI 1 using SNAI 1 siRNA and shRNA Plasmids (h) from Santa Cruz Biotechnology. Please note that these videos are representative and steps or experiment specific processes must be kept in mind to expect desired results.

We haven't found any additional videos for this experiment / product combination yet.

Outsource your experiment

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

  Outsource experiment
Become shareholder Discussions About us Contact Privacy Terms