Nucleic acid releasing agent NF-3,RPA

Nucleic acid releasing agent NF-2/NF-3/NF-4 can directly cleave and release sample nucleic acid (DNA/RNA) without nucleic acid extraction. The supernatant of the mixed solution is directly used as a template for nucleic acid PCR amplification or isothermal amplification, and the buffer has specific protection for nucleic acid amplification enzymes.

Nucleic acid releasing agent NF-2/NF-3/NF-4 is suitable for efficient inactivation of cells (eukaryotes, insects, etc.) and pathogens (viruses, mycoplasma, chlamydia, bacteria, etc.) in all clinical samples (saliva, fresh whole blood, serum, alveolar lavage fluid, urine, tissue, environment, etc.).

The mixed solution does not need to extract nucleic acid, and the supernatant can be directly taken as a template for PCR or isothermal amplification detection. The amplification efficiency was 2 ~ 4 CT values lower than that after nucleic acid extraction, and the sensitivity of fluorescence PCR detection after nucleic acid release was up to 5 cells, about 500~1000copies/mL.

The pathogen nucleic acid released can be preserved at room temperature for a long time, in which the RNA released at room temperature does not degrade for 7 days, and the DNA released at room temperature does not degrade for 6 months.

Release steps of different samples

 1. Serum, plasma, saliva, sputum, urine, non-PBS swabs, non-guanidine salt collection fluid, cell culture fluid and other liquid samples.

1 the release agent 150 μ L was placed in the 1.5mL centrifuge tube, the liquid sample 50 μ L was added, the liquid sample was blown and mixed for 2-5 times, and the effect was ≥ 2min at room temperature (22-25 ℃).

2 if it is fresh whole blood, take 10 μ L whole blood, add 100 μ L release agent, repeatedly blow and mix for 2-5 times, the effect is ≥ 2min at room temperature (22-25 ℃).

2.EDTA anticoagulant samples, complete hemolysis samples, concentrated sputum, ascites samples, high concentration PBS solution samples.

Need to first dilute with DEPC water 10 times, and then release according to the liquid sample: release agent = 1:3, such as take the sample diluted 10 times 50 μ L, and then add the release agent 150 μ L, after mixing, place the static effect ≥ 2min at room temperature.

3. Tissue solid sample.

The release agent 300 μ L was placed in the 1.5mL centrifuge tube, and then the tissue sample with sufficient soybean size was added, and the tissue sample was repeatedly blown and mixed for 3-5 times. The effect was ≥ 5min at room temperature (22-25 ℃).

Note: the supernatant of normal saline tissue homogenate can also be added and operated according to the liquid sample.

Freeze-dried amplification sampling of releasing agent NF-3/NF-4.

25 ~ 50 μ L of the supernatant of the sampling tube was directly taken as freeze-dried RPA, PCR or LAMP amplification template for RPA, PCR or LAMP detection.
Note: the volume of the template should be the total volume required by the freeze-drying system, that is, the sample tube supernatant of 25 μ L freeze-drying reaction system is 23 μ L, and that of 50 μ L freeze-drying reaction system is 45 μ L (exactly one drop).