Molecular marker TCONS_00016233, kit and application
A technology of molecular markers and kits, applied in the field of molecular markers for predicting sepsis complicated with acute kidney injury, early diagnosis, and can solve problems such as inaccurate reflection of renal function
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Embodiment 1
[0020] Example 1: Screening for molecular markers associated with sepsis complicated by acute kidney injury
[0021] 1. Sample Collection
[0022] Blood samples were collected from healthy people, patients with sepsis without acute kidney injury, and patients with sepsis complicated with acute kidney injury.
[0023] 2. RNA sample preparation and quality analysis
[0024] Use the Trizol of Kangwei Century Company to extract total RNA, and the specific operation steps are as follows:
[0025] 1) Take 2ml of whole blood collected in a test tube treated with sodium citrate, and put it into an enzyme-free centrifuge tube;
[0026] 2) Collect plasma: Centrifuge at 3000rpm for 10min, carefully suck the supernatant (plasma) from the top of the sample and put it into another enzyme-free centrifuge tube;
[0027] 3) Take 250 μL of plasma liquid, transfer it to a 1.5ml centrifuge tube, add 750 μL of TRIzol reagent, shake the tube body vigorously by hand until mixed.
[0028] 4) Afte...
Embodiment 2
[0043] Embodiment 2: RT-PCR verification differential expression
[0044]
[0045]
[0046] 1. According to the detection results of high-throughput sequencing, select for RT-PCR verification. According to the sample collection method in Example 1, blood samples from healthy people, sepsis complicated with acute kidney injury, and sepsis without acute kidney injury were selected.
[0047] 2. RNA extraction steps are the same as in Example 1
[0048] 3. Reverse transcription: use the reverse transcription kit from ThermoFish Company to operate, and the specific steps are as follows:
[0049] 1) Add the reagents used, store on ice, and conduct related experiments according to the table below.
[0050] 2) Store at -20°C for later use.
[0051] 4. RT-PCR amplification
[0052] 1) Primer design
[0053] The primers for PCR amplification were designed according to the gene coding sequence of TCONS_00016233 and Gapdh, and were synthesized by Guangzhou Ruqi Biotechnology Co.,...
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