Enterobacter sakazakii fluorescence real-time PCR test

Specification: 96 T / box

Uses: Real-time fluorescent PCR in vitro detection of enterobacter sakazakii.

Detection principle: The kit contains enterobacter sakazakii-specific primers. When the sample contains enterobacter sakazakii, the enterobacter sakazakii DNA extracted after pre-enrichment is exponentially amplified by PCR. During the reaction, the enterobacter sakazakii-specific fluorescent probe hybridizes with the target nucleic acid and is simultaneously taq enzyme Hydrolysis to separate the fluorescent group from the quenching group on the probe: so that the presence of enterobacter sakazakii can be judged in real time by the fluorescence increment.

Product and composition:

Storage conditions and expiration date of the kit:
1. The kit is stored at 2 -8℃.
2. The validity period is 12 months.

Applicable instruments:
ABI7500, 9600 series, MJ opticon2, Bio-Rad and other fluorescent quantitative PCR instruments are available

Experimental method:
1. Take 1ml of the enrichment solution into a 1.5m1 sterile centrifuge tube, centrifuge at 10.000rpm for 5min, and discard the supernatant; use the classic phenol chloroform extraction method to test the extracted template or store it at 20C until it is tested.
2. Place the PCR reaction solution at room temperature to equilibrate, take the Taq enzyme after melting, and add the Taq enzyme at 2∪/tube, that is, each test reaction system is prepared as: PCR reaction solution 22.5u1+0.4ulTaq enzyme.
3. Add the template: Thaw the nucleic acid extract stored at -20°C at room temperature and centrifuge at 13,000 rpm for 5 minutes. The negative and positive controls are thawed at room temperature before use. In each PCR reaction tube, add 2 ul of the template or negative and positive controls processed in step 1, cover the tube, place on the PCR instrument, and record the corresponding sample number.
4. On-board amplification and detection area: the reaction conditions are 95C: 3min, 1 cycle; 95C: 5sec, 60C: 40sec (signal acquisition), 40 cycles. The reaction system was set to 25u1.
For the multi-channel fluorescence PCR instrument, the signal is set to Fam fluorescein.

Results judgment:
Please refer to the manual for details.

Precautions for using the kit:
1. This kit is only used for in vitro testing and research purposes, please read the full text of this manual carefully before starting to use it.
2. The experiment must be strictly partitioned: pre-PCR preparation area-prepare experimental reagents; sample processing area-to be tested samples, template processing; detection area-PCR amplification, detection.
3. For the relevant laboratory management standards, please refer to the implementation of the management standards for gene amplification laboratories issued by relevant national authorities.