Fluorescent Probe PCR Kit

The nucleic acid extraction of the sample to be tested using fluorescent probe PCR kit can be carried out using virus RNA extraction kit or automated nucleic acid extraction instrument. Please refer to the relevant instructions for specific extraction methods; The instructions for the PCR identification kit for positive and negative samples of Artemisia argyi do not require extraction and can be used directly.

Characteristics of Fluorescent Probe PCR Kit:

This product is a highly sensitive fluorescent quantitative PCR product developed based on primers designed for conservative regions.

The principle is mediated by a pair of primers, which can rapidly amplify specific gene (DNA) fragments in vitro in animals and plants. After n thermal cycles of amplification, the number of specific genes contained in the amplification product is (1+E) n (0) of the original template number

Transportation: Low temperature

Save: Negative 20 degrees

Validity period: One year

Delivery time: 2-3 weeks

Notes: 1. Basic program; 2. Amplification temperature and extension temperature; 3 reaction time; 4 cycles; Preparation of PCR reaction solution; The basic principles of 6PCR technology; The reaction kinetics of 7PCR; 8PCR amplification product; 9 PCR reaction system and reaction conditions.

Usage:

1、 Dilute the standard curve sample (using 6 10 fold dilutions of 10E2-10E7 copies/μ L as an example). Due to the high concentration of the standard substance, the following dilution operations must be carried out in a separate area and must not contaminate the sample or other components of this kit.

1. Mark 6 centrifuge tubes, namely 7, 6, 5, 4, 3, and 2.

2. Add 45 μ L of fluorescent PCR template dilution solution using a core gun tip, the same applies below.

3. Add 5 μ L of positive control (concentration of 1 × 10E8 copy/μ L, provided by the reagent kit) to tube 7, shake thoroughly for 1 minute, and obtain a standard curve sample of 1 × 10E7 copy/μ L. Put it on ice for later use.

4. Change the gun head and add 5 μ L of 1 × 10E7 copy/μ L positive control (diluted in the previous step) to tube 6. Shake thoroughly for 1 minute to obtain a standard curve sample of 1 × 10E6 copy/μ L. Put it on ice for later use.

5. Change the gun head and add 5 μ L of 1 × 10E6 copy/μ L positive control (diluted in the previous step) to tube 5. Shake thoroughly for 1 minute to obtain a standard curve sample of 1 × 10E5 copy/μ L. Put it on ice for later use.

6. Repeat the above operation until obtaining standard curve samples with 6 dilutions. Put it on ice for later use.

2、 Preparation of Sample DNA

7. Purify the DNA of the sample using a self selected method. This product is compatible with the vast majority of nucleic acid purification products on the market.

If there are N samples, N+2 sample extractions are required, with the extra one used as a positive control tube for sample preparation and the other as a negative control tube for sample preparation.

3、 Set up qPCR reaction (20 μ L system, conducted in the sample preparation room)

9. If quantitative analysis is performed and only one replicate is performed, label N+9 PCR tubes, including N+2 for the N+2 samples obtained in the previous step, 1 for PCR negative control (using water as a template), and 6 for the standard curve. If qualitative analysis is performed and only one repetition is made, label N+4 PCR tubes, including N+2 for the N+2 samples obtained in the previous step, 1 for PCR negative control (using water as a template), and 1 for PCR positive control.

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　　Fluorescent Probe PCR KitNotes:

① The order of adding reagents should be consistent to ensure that the incubation time for all reaction plate wells is the same.

② Use clean plastic containers to prepare detergent.

③ Incubate according to the time, amount, and sequence indicated in the instructions of the chicken hemoglobin (HB) ELISA detection kit.

④ Substrate A should evaporate and avoid opening the lid for a long time. Substrate B is sensitive to light and should be avoided from prolonged exposure to light. Avoid contact with hands, toxic. After the experiment is completed, the OD value should be read immediately.

⑤ When washing the enzyme-linked immunosorbent assay (ELISA) plate, it should be thoroughly dried. Do not directly put the absorbent paper into the ELISA reaction well to absorb water.

⑥ Use disposable suction tips to avoid cross contamination, and avoid using sample dispensers with metal parts when suctioning termination solution and substrates A and B.

⑦ The Flat noodles not used in the experiment shall be immediately put back into the packaging bag and sealed for storage to avoid deterioration.

⑧ Reagents should be stored according to the label instructions and returned to room temperature before use. The diluted standard should be discarded and cannot be stored.

⑨ Other unused reagents should be packaged or covered. Do not mix reagents of different batches. Use before shelf life.