Related serum microribonucleic acid marker for human severe preeclampsia and application of marker

A technology of preeclampsia and markers, applied in the fields of genetic engineering and clinical medicine, can solve problems that have not received corresponding attention

Active Publication Date: 2013-07-17
夏彦恺
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the application of serum miRNAs in the early diagnosis and monitoring of severe preeclampsia has not received corresponding attention. If stable specific serum miRNAs related to the onset of severe preeclampsia can be found as biomarkers, and the diagnosis and development of corresponding diseases, The monitoring kit is not only in an international leading position in this field, but also can create remarkable economic benefits, and it will also be a strong impetus to the prevention and treatment of pregnancy-induced hypertension in my country

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  • Related serum microribonucleic acid marker for human severe preeclampsia and application of marker
  • Related serum microribonucleic acid marker for human severe preeclampsia and application of marker
  • Related serum microribonucleic acid marker for human severe preeclampsia and application of marker

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Embodiment 1

[0076] Embodiment 1 Research Object Selection and Grouping Basis

[0077] From July 2009 to October 2011, the inventor collected the blood samples of severe preeclampsia patients and healthy control pregnant women who met the requirements from the Affiliated Hospital of Nanjing Medical University three times. 100 healthy controls (average age: 27.63±3.25 days) and 100 patients with severe preeclampsia (average age: 28.56±4.15 days) were used as experimental subjects for Real-time PCR detection of miRNA expression. Specific sample classification criteria are as follows:

[0078] Group A: healthy control group (n=100, 20 people for microarray screening, 40 people for first-stage verification, 40 people for independent population verification):

[0079] 1. No other major systemic diseases.

[0080]Group B: Severe preeclampsia group (n=100, 20 people for microarray screening, 40 people for phase one verification, 40 people for independent population verification):

[0081] 1. C...

Embodiment 2

[0083] Example 2 TaqMan miRNA array screening

[0084] Preparation of cDNA samples: a) Take 100 μl of serum; b) Add 900 μl of Trizol, shake and mix, centrifuge at 12,000 rpm at 4°C for 15 minutes, and discard the waste liquid in the lower layer; c) Add 1.5 times the volume of supernatant, shake and mix, transfer To the spin column, centrifuge at 12000rpm for 15 seconds, discard the lower waste liquid; d) add 700μl RWT buffer to the spin column, centrifuge at 10000rpm for 15 seconds, discard the lower waste liquid. e) Add 500 μl of RPE buffer to the spin column, centrifuge at 10,000 rpm for 15 seconds, and discard the lower layer. f) Repeat e. g) Add the spin column to a new 2ml tube and centrifuge at 10,000rpm for 1 minute to remove the RPE buffer. h) Add 50 μl of DEPC-treated water to the column and collect RNA by centrifugation at 12000 rpm. i) Then cDNA is obtained by RNA reverse transcription reaction. The reverse transcription reaction system included 4 μl 5×AMV buffe...

Embodiment 3

[0095] Embodiment 3Real-time PCR method measures serum miRNA expression

[0096] Primers (Table 1) were designed for quantitative Real-time PCR detection of each miRNAs in the serum of 80 healthy controls and 80 patients with severe preeclampsia.

[0097] (1) Preparation of cDNA samples: a) Take 100 μl of serum; b) Add 900 μl of Trizol, shake and mix, centrifuge at 12,000 rpm at 4°C for 15 minutes, and discard the waste liquid in the lower layer; c) Add 1.5 times the volume of supernatant in absolute ethanol, shake and mix Mix well, transfer to the spin column, centrifuge at 12000rpm for 15 seconds, discard the lower waste liquid; d) add 700μl RWT buffer to the spin column, centrifuge at 10000rpm for 15 seconds, discard the lower waste liquid. e) Add 500 μl of RPE buffer to the spin column, centrifuge at 10,000 rpm for 15 seconds, and discard the lower layer. f) Repeat e. g) Add the spin column to a new 2ml tube and centrifuge at 10,000rpm for 1 minute to remove the RPE buff...

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Abstract

The invention belongs to the fields of genetic engineering and clinical medicine, and discloses a related serum microribonucleic acid marker for human severe preeclampsia and application of the marker. The marker is selected from more of hsa-miR-193a-5p, hsa-miR-424 and hsa-miR-10a. The marker has the specificity and sensitivity on a sufferer with severe preeclampsia, and can be used for preparing a diagnosis or monitoring kit for severe preeclampsia.

Description

field of invention [0001] The invention belongs to the fields of genetic engineering and clinical medicine, and relates to serum microRNA (miRNA) markers related to the occurrence of human severe preeclampsia and applications thereof. Background technique [0002] The incidence rate of hypertensive disorders during pregnancy in my country is about 9.4%, and it is 7% to 12% in foreign reports. It is an important cause of maternal and perinatal deaths. Hypertensive disorders of pregnancy include gestational hypertension, preeclampsia, eclampsia, chronic hypertension with preeclampsia, and chronic hypertension. Preeclampsia (PE) is a multisystem disease unique to pregnancy, with an incidence rate of 1% to 5%. The main clinical features are hypertension, proteinuria, and edema after 20 weeks of pregnancy. Preeclampsia is divided into mild and severe according to the severity of the disease. Mild preeclampsia refers to blood pressure ≥ 140 / 90mmHg, urine protein (+) or urine pro...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12N15/11C12Q1/68
Inventor 夏彦恺吴炜吕述彦傅广波陆春城王心如
Owner 夏彦恺
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