Molecular marker and kit for early diagnosis and prediction of septicopyemia complicated by acute kidney injury and application thereof
A technology of acute kidney injury and molecular markers, which can be used in disease diagnosis, analytical materials, measuring devices, etc., 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] Collect healthy people, sepsis non-acute kidney injury, sepsis complicated with acute kidney injury patients
[0023] blood sample.
[0024] 2. RNA sample preparation and quality analysis
[0025] Use the Trizol of Kangwei Century Company to extract total RNA, and the specific operation steps are as follows:
[0026] 1) Take 2 mL of whole blood collected in a sodium citrate-treated test tube and put it into an enzyme-free centrifuge tube;
[0027] 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;
[0028] 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.
[0029] 4) After homogenization, the samples were ...
Embodiment 2
[0045] Embodiment 2: RT-PCR verification differential expression
[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 of ThermoFish Company to operate.
[0049]
[0050] 4. RT-PCR amplification
[0051] 1) Primer design
[0052] The primers for PCR amplification were designed according to the gene coding sequence of TCONS_00024536 and Gapdh, and were synthesized by Shanghai Sangon Bioengineering Technology Service Co., Ltd. The specific primer sequences are as follows:
[0053] TCONS_00024536:
[0054] F: 5'-TAGGAAGGGCTGTTGACTGG-3'
[0055] R: 5'-CTGGGAGCTGGATTCAGAAG-3'
[0056] Gapdh gene:
[00...
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