NuProZol Reagent
Trizol domestic alternative, high yield and high purity
One-click copy of information
Instruction manual- Product Description
- Composition
- Precautions
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Transportation and Storage Conditions: NuProZol Reagent should be stored and transported at 2–8°C, with a shelf life of one year.
Product Overview
NuProZol Reagent is designed for the rapid extraction of high‑quality total RNA from cells and animal tissue samples. This reagent contains guanidinium isothiocyanate and phenol, which efficiently lyse biological samples and inactivate RNases, ensuring the integrity of the extracted RNA. NuProZol Reagent rapidly lyses the sample; following extraction with chloroform or a chloroform substitute, the mixture separates into an upper aqueous phase (containing RNA), an interphase, and a red organic phase. Total RNA of high yield and purity is then recovered by precipitation with isopropanol and washing with ethanol. The resulting total RNA can be directly used in molecular biology applications such as RT‑PCR, qRT‑PCR, Northern blotting, and high‑throughput sequencing.
Preparations
1. This reagent contains phenol. During the final step, add RNase‑free water to dissolve the precipitate. Perform all steps in a fume hood and wear appropriate protective equipment, such as protective clothing, gloves, eye protection, and a face shield. If the reagent comes into contact with the eyes, rinse immediately with copious amounts of water and seek medical attention. If it contacts the skin, wash thoroughly with a detergent and plenty of water; if irritation persists, seek medical care.
2. Reagents to be provided by the user: chloroform or a chloroform substitute (catalog no.: 400-301), isopropanol (newly opened or for RNA extraction only), 75% ethanol (prepared with DEPC‑treated water), and RNase‑free water or DEPC‑treated water.
3. Pre-run the 4°C centrifuge to bring its temperature down to 4°C.
Operating steps
1. Homogenization treatment
a. Animal tissue
Homogenize the tissue by adding 1 ml of NuProZol Reagent lysis buffer per 50–100 mg of tissue. The volume of the tissue sample should not exceed 10% of the volume of the NuProZol Reagent. Grinding should be as thorough and rapid as possible, and performed at low temperature to prevent RNA degradation.
b. Adherent cells
Discard the culture medium, then directly add 1 ml of NuProZol Reagent lysis buffer to a 3.5‑cm‑diameter culture plate, ensuring complete coverage, and pipette up and down several times to lyse the cells. Determine the required volume of NuProZol Reagent based on the surface area of the culture plate rather than the number of cells (1 ml per 10 cm²). Insufficient NuProZol Reagent can lead to DNA contamination.
c. Suspended-growth cells
Centrifuge to pellet the cells, then carefully discard the supernatant. Add 1 ml of NuProZol Reagent per 5–10 × 10^6 animal cells. Pipette up and down repeatedly in the NuProZol Reagent to lyse the cells.
2. Vigorously vortex the homogenized sample 1–3 times, then allow it to stand at room temperature for 5 minutes to ensure complete disruption of ribonucleoprotein complexes.
3. Add 0.2 mL of chloroform or a chloroform substitute (Catalog No.: 400-301). Secure the sample tube cap, shake vigorously for 15 seconds, and let stand at room temperature for 3 minutes.
4. Centrifuge at 12,000×g for 15 minutes at 4°C; the sample will separate into three layers: an organic phase at the bottom, an interphase, and a colorless aqueous phase on top, with RNA residing in the aqueous phase. Transfer approximately 500 μl of the aqueous phase to a new tube (avoiding the interphase).
5. Add an equal volume of isopropanol, invert and mix thoroughly, then let stand at room temperature for 10 minutes. Centrifuge at 12,000 × g and 4°C for 10 minutes; a white precipitate will typically be visible. Carefully discard the supernatant.
6. Add 1 mL of 75% ethanol (prepared with DEPC‑treated water or RNase‑free water). Gently flick the bottom of the tube to resuspend the pellet, and invert the tube several times.
7. Centrifuge at 4°C at 12,000×g for 5 minutes, then carefully discard the supernatant. Note: The small amount of residual liquid may be briefly centrifuged and then aspirated with a pipette tip; be careful not to remove the pellet.
8. In a clean bench, allow the precipitate to air-dry at room temperature for 2–5 minutes. Then dissolve the pellet in 30–100 μL of DEPC‑treated water or RNase‑free water, vortexing at room temperature or repeatedly pipetting up and down to ensure complete dissolution of the RNA.
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Product Composition
Product Number Component Specifications 400-300 NuProZol Reagent 100 ml -
Precautions
1. Use high-quality centrifuge tubes and consumables to prevent tube breakage or reagent spillage during experiments.
2. To ensure extraction quality, it is recommended to use RNase-free reagents and consumables throughout the entire process, particularly during the RNA elution step.
3. After homogenizing the sample with NuProZol Reagent, if the assay is not performed immediately, it can be stored at −70°C for up to one month prior to the addition of chloroform. Store in…
RNA precipitated in 75% ethanol can be stored at 2–8°C for one week and at −20°C for up to one year. Due to its relatively short half-life, RNA is prone to degradation; therefore, it is recommended to perform downstream applications—such as reverse transcription into cDNA or Northern blotting—as soon as possible after extraction.
Keywords
GOONIE
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Goonie Biotechnology (Guangzhou) Co., Ltd.
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