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hsa-miR-195-3p inhibitor

CAS No. :

MCE 站：hsa-miR-195-3p inhibitor

产品活性：hsa-miR-195-3p inhibitor 是一种全链经过甲氧基修饰的成熟miRNA的互补单链。miRNA inhibitor通过特异性结合成熟miRNA，阻止miRNA与其靶基因的互补配对，从而抑制miRNA的功能，可用于miRNA 功能缺失性（loss-of-function）研究。

研究领域：Epigenetics

作用靶点：MicroRNA

In Vitro:

1. miRNA Resuspension

1.1 Briefly centrifuge the tube to ensure that the dried miRNA is at the bottom of the tube.

1.2 Resuspend the miRNA using nuclease free water to generate 20 μM stock solution.

For 5 nmol miRNA: add 250 μL nuclease free water.

For 20 nmol miRNA: add 1000 μL nuclease free water.

1.3 Aliquot miRNAs into one or more tubes to limit the number of freeze-thaw cycles (<5).

1.4 Store at or below -20°C or -80°C in a non-frost-free freezer until use.

2. Prepare cells

2.1 Inoculate cells in advance for cell transfection. The viability ａnd general health of cells prior to transfection significantly affect transfection result.

3. Transfection

3.1 Prepare transfection mix A ａnd B.

For per well of a 6-well plate: A: 240 μL serum-free medium + 10 μL miRNA; B: 230 μL serum-free medium + 20 μL HY-K2017 siRNA/miRNA Transfection Reagent.

For per well of a 12-well plate: A: 95 μL serum-free medium + 5 μL miRNA; B: 90 μL serum-free medium + 10 μL HY-K2017 siRNA/miRNA Transfection Reagent.

For per well of a 24-well plate: A: 47.5 μL serum-free medium + 2.5 μL miRNA; B: 45 μL serum-free medium + 5 μL HY-K2017 siRNA/miRNA Transfection Reagent.

For per well of a 96-well plate: A: 24.5 μL serum-free medium + 0.5 μL miRNA; B: 24 μL serum-free medium + 1 μL HY-K2017 siRNA/miRNA Transfection Reagent.

Note: The recommended working concentration is 100nM for miRNA inhibitors. miRNA function can vary greatly, depending on the miRNA, the cell line, ａnd the chosen analysis method. To determine the concentration that provides optimal results, optimization experiments using varying mimic/inhibitor concentrations should be performed. The optimized range suggests changing the miRNA concentration in the range of 20 to 500nM.

If other transfection reagents are used, the amount of transfection reagent needs to be adjusted according to the specific situation.

3.2 Mix A ａnd B gently. Incubate at room temperature for 15 minutes.

3.3 Remove culture medium from cells, wash with PBS.

3.4 Add transfection mix (A+B) to cells.

For per well of a 6-well plate: add 1500 μL serum-free medium, then add 500 μL of the transfection mix (A+B) to the well, ａnd mix well.

For per well of a 12-well plate: add 800 μL serum-free medium, then add 200 μL of the transfection mix (A+B) to the well, ａnd mix well.

For per well of a 24-well plate: add 400 μL serum-free medium, then add 100 μL of the transfection mix (A+B) to the well, ａnd mix well.

For per well of a 96-well plate: add 50 μL serum-free medium, then add 50 μL of the transfection mix (A+B) to the well, ａnd mix well.

3.5 Incubate cells for 13 days at 37°C. Then, analyze transfected cells. The medium can be replaced with fresh serum-containing medium after 6 hours if necessary.

Note: Antibiotics can increase toxicity ａnd should be omitted during transfection. Culture medium containing polyanions such as heparin, heparin sulfate or dextran sulfate can inhibit transfection.

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