Complex Plant Total RNA (più) Small Amount Extraction Kit

1. Componenti del kit di reagenti

2. Magazzinaggio

This reagent kit can be stored at room temperature (15-25℃) in a dry environment and is stable for 12 mesi.

3. Istruzioni per l'uso del kit di reagenti

3.1 Questo kit è destinato a scopi di ricerca in biologia molecolare e non deve essere utilizzato per la diagnosi o il trattamento di malattie.

3.2 Alcuni componenti del kit contengono sostanze irritanti; è opportuno prendere le dovute precauzioni (come indossare indumenti protettivi e occhiali protettivi).

3.3 L'utilizzo di questo kit richiede apparecchiature aggiuntive come una centrifuga ad alta velocità, bagnomaria (bagno di metallo), miscelatore a vortice, etanolo anidro, azoto liquido, cloroformio, acqua deionizzata sterile, e tubi EP.

4. Introduzione al kit di reagenti

This RNA purification reagent kit provides a fast and effective purification of plant total RNA containing polysaccharides, lipids, polyphenols, and other components. It is suitable for most complex plant tissues. Generalmente, lipid-rich plant tissues contain a high content of lipid compounds, which significantly affect RNA extraction efficiency. This kit utilizes exclusive inhibitor removal columns to effectively eliminate lipids from plant samples, as well as to remove DNA contamination. If the experiment is sensitive to DNA, it is recommended to use intron-spanning primers for downstream experiments.

The RNA rapid purification reagent kit can extract plant total RNA (including nuclear RNA and cytoplasmic RNA) entro 1 ora. The extracted RNA can be directly used for RT-PCR, Northern blotting, eccetera. The entire purification process does not require toxic reagents such as chloroform, making the RNA purification reagent kit suitable for various other samples.

5. Principi e procedure sperimentali

6. Processo di estrazione

Precauzioni prima di iniziare l'esperimento:

UN. Lavare Respingente 1: Prior to use, add the specified amount of absolute ethanol as indicated on the reagent bottle label. Check the label to confirm the addition of absolute ethanol.

B. Il tampone di eluizione è un 0.1x soluzione TE with a minimal amount of EDTA. If EDTA may affect subsequent experiments, it is recommended to replace the elution buffer with sterile deionized water.

C. RNA Extraction Buffer I contains a small amount of phenol, which may precipitate. Prima dell'uso, mix thoroughly by warming in a water bath; after use, store away from light.

1. Elaborazione del campione:

UN. Raccolta e stoccaggio dei materiali:

Se i materiali appena raccolti non possono essere utilizzati immediatamente, place them in liquid nitrogen and store them at -80℃.

B. Whenever possible, collect fresh materials as they contain fewer polysaccharides and polyphenols.

2. Grind approximately 100 mg of fresh samples or not more than 20 mg of dry material with liquid nitrogen.

3. Aggiungere 600μl of RNA Extraction Buffer I, ensuring there are no tissue clumps in the ground sample. Tissue clumps are difficult to lyse and can reduce RNA yield.

4. Vortex for 30 S.

5. Transfer the lysate to aninhibitor removal purification column, centrifugare a 12,000 giri al minuto per 5 min.

(Nota: Lipid-rich plant materials may contain many lipid compounds at this step, which can affect RNA extraction. Remove these substances during this step. If buffer residue remains in the inhibitor removal purification column during centrifugation, extend centrifugation time appropriately.)

6. Transfer the obtained supernatant to a DNA removal column, centrifugare a 12,000 rpm for 30s, and collect the filtrate (Nota: RNA is present in the filtrate).
7. Aggiungere 250μl of absolute ethanol, mescolare pipettando. If there is a small amount of precipitation, it does not affect subsequent experiments. Transfer the liquid to an RNA purification column, centrifugare a 12,000 rpm for 30s, discard the flow-through.
8. Aggiungere 700μl of inhibitor removal buffer, centrifugare a 12,000 rpm for 30s, discard the flow-through.
9. Aggiungere 700ml di lavarerespingente 1 to the RNA purification column, centrifugare a 12,000 rpm for 30s, discard the flow-through.
10. Aggiungere 500ml di lavare respingente 1 to the RNA purification column, centrifugare a 12,000 giri al minuto per 3 min, discard the flow-through.
11. Place the RNA purification column into a new 1.5ml centrifuge tube, air-dry the membrane at room temperature for 2 minuti.

(Nota: Confirm that absolute ethanol has been added to lavare respingente 1. The presence of ethanol has a serious impact on subsequent experiments, so drying the membrane is crucial. Dopo la centrifugazione, ensure no ethanol is present before elution, then discard the waste and collection tube. After washing with wash buffer 1, the membrane on the RNA purification column should have only a slight color. Dopo la centrifugazione, carefully remove the RNA purification column, ensuring it does not touch the collection tube to avoid ethanol interference.)

12. Aerially pipette 50-100ml di tampone di eluizione sulla membrana, centrifugare a 12,000 giri al minuto per 1 min, and collect the RNA solution.

(Nota: Eluting RNA with 50 μl of elution buffer can increase RNA concentration but reduces total RNA yield.)