Transport temperature: normal temperature transport.

Product Introduction:

This reagent is a general total RNA extraction reagent, which is suitable for the total RNA extraction of animal and plant cells or tissues and bacteria, and can effectively prevent the degradation of RNA during the extraction process. The obtained RNA can be directly used in Northern hybridization, purified mRNA, in vitro translation, RNase protection assay, RT-PCR, cDNA cloning and other operations. The reagent can be used for 100 total RNA extractions (10cm2 cells or 100mg tissue).

Operating instructions:

Bring freshly opened or dedicated chloroform, isopropyl alcohol, 75% ethanol, DEPC treated water.

1. Cell lysis or tissue homogenate:

1) Adherent cells: Soak up the culture solution, add 1mL TriQuick Reagent for every 10cm2 (6-well plate hole or 35mm plate) cells, make it cover the cultured cells, and then blow with a straw or sampler for 2 to 3 times, the cells should be completely lysed, and then transferred to the centrifuge tube.

2) Suspension cells: Centrifuge the cells, suck up the fluid, and add 1mL of TriQuick ReagentL for every 5 to 10 million (5×106-1×107) animal or yeast cells, or 10 million (1×107) bacteria. Blow it with a straw or injector to break it completely. Certain yeasts and bacteria, such as insufficient cracking, can be homogenized in a homogenizer to ensure complete cracking. Transfer to centrifuge tube.

3) Tissue: First cut the tissue into small pieces and put them into the glass homogenizer. The frozen tissue can be homogenized in a mortar. 1mL TriQuick ReagentL is added to every 50-100mg tissue and homogenized until completely cracked. Transfer to centrifuge tube.

The pyrolysis product should be a clear transparent viscous liquid. Leave for 5 minutes at room temperature. For tissue samples rich in impurities such as polysaccharide and protein, there will still be insoluble substances after homogenization, which can be centrifuged at 12000g and 4℃ for 10min, and then absorb the supernatant into a new centrifuge tube.

2. Separation: Add 0.2 times the volume of chloroform (0.2ml of chloroform for 1mL TriQuick Reagent) into the centrifuge tube equipped with lysate, fully oscillate and mix for 30s on the oscillator, and place at room temperature for 2-3min. 12000g, centrifuge at 4℃ for 10min, and then absorb the upper water phase containing total RNA into a new centrifuge tube, about 0.5-0.6mL per milliliter of TriQuick Reagent can be absorbed. The organic phase and mesosphere contain DNA and proteins and should be avoided.

3. Precipitation: Add 0.5mL isopropyl alcohol per ml of TriQuick Reagent, reverse mixing several times, and precipitate at room temperature for 10min. At 12000g, centrifuged at 4℃ for 10min, RNA precipitation was observed at the bottom of the tube. Discard the supernatant, add 1mL 75% ethanol per milliliter of TriQuick Reagent, and gently mix upside down to clean the RNA precipitation. Centrifuge 12000g at 4℃ for 2min, discard the liquid, and be careful not to discard the RNA precipitate. Dry upside down at room temperature for 5~10min.

4. Dissolve: Add proper amount of DEPC treated water to dissolve RNA precipitation. Store at -80℃.

Note:

1. Vessels, centrifugal tubes and suction tubes used in the RNA extraction process should be guaranteed to be pollution-free and RNA-free. Care should be taken during operation to prevent contamination of the sample by exogenous RNase leading to degradation of the RNA sample.

2, the reagent contains phenol and other harmful substances, pay attention to personal protection."

Note：Product information may be optimized and upgraded. Please refer to the actual label information for accuracy.