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Kit for screening or early diagnosis of gestational diabetic huge high-risk pregnant women

A kit and reagent technology, applied in DNA/RNA fragmentation, microbial determination/examination, recombinant DNA technology, etc., to achieve the effect of reducing adverse pregnancy outcomes, far-reaching clinical significance and good promotion and specificity

Active Publication Date: 2022-07-05
NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no relevant research report on exosomal lncRNA as a biomarker for macrosomia

Method used

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  • Kit for screening or early diagnosis of gestational diabetic huge high-risk pregnant women
  • Kit for screening or early diagnosis of gestational diabetic huge high-risk pregnant women

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Collection of samples and arrangement of sample data

[0024] The applicant collected a large number of peripheral blood samples of pregnant women at 24-28 weeks of gestation from Nanjing Maternal and Child Health Hospital from January to December 2020 (samples used for research were collected during the same period, and the sampling, packaging and storage conditions were uniform), By sorting out the sample data, the applicant selected 80 samples that met the following standards as experimental samples.

[0025] 1. Pregnant women who were 24-28 gestational weeks at the time of blood collection and diagnosed with GDM by OGTT;

[0026] 2. Pregnant women whose newborns weighed more than 4000g after the full-term study subjects were defined as the experimental group (40 cases);

[0027] 3. The weight of the newborns born to the above-mentioned subjects after full-term should not exceed 4000g. The pregnant women who matched the age, BMI and gestational age of the ...

Embodiment 2

[0029] Example 2 Separation of exosomes in serum, extraction and purification of RNA in exosomes

[0030] 1. Separation of exosomes in serum: Add 100ul of Total ExosomeIsolation Reagent to 500ul of each serum sample, mix by vortexing, and react at 4°C for 30min. Centrifuge at 10000g for 10min at room temperature. There are exosomes at the bottom of the EP tube, resuspend the exosomes with 200uI of PBS (select a commercialized exosome isolation kit);

[0031] 2. Extraction and purification of RNA from exosomes (select a commercialized exosome isolation and purification RNA kit):

[0032] 2.1 Extraction of RNA from exosomes: Add 200ul of 2X Denaturing Solution and mix, incubate on ice for 5min, then add 400ul of acid-Phenol:Chloroform, and vortex for 60s. Centrifuge at 12000g for 10min at room temperature, the supernatant contains RNA;

[0033] 2.2 RNA purification: Pipette 300ul of supernatant into an enzyme-free EP tube, add 375ul of absolute ethanol, and mix the two. The ...

Embodiment 3

[0034] Embodiment 3 adopts the method of reverse transcription and real-time quantification to detect LncRNA AC015961.2

[0035] 1. Preparation of LncRNA AC015961.2 kit (50 reactions) for screening or early diagnosis of pregnant women with high risk of GDM macrosomia, the kit includes:

[0036] 1.1 Enzyme-free water 10ml

[0037] 1.2 Random reverse transcription primer (1uM) 50ul

[0038] 1.3 5X reverse transcription buffer 200ul

[0039] 1.4 triphosphate base deoxynucleotide (10mM) 100ul

[0040]1.5 RNase inhibitor (40M / ul) 50ul

[0041] 1.6MMLV reverse transcriptase (200M / ul) 50ul

[0042] 1.7SYBR Premix Ex Taq 500ul

[0043] 1.8LncRNA AC015961.2 Real-time PCR specific primer (10uM) 30ul

[0044] The forward primer is 5'-CTGCAAGCACCAAGAACGAAA-3' (SEQ ID NO.2)

[0045] The negative primer is 5'-GGAGAAAGTCCCCCATCCG-3' (SEQ ID NO.3)

[0046] 1.9U6snRNA internal reference specific PCR primer (10uM) 30ul

[0047] The forward primer is 5'-ATTGGAACGATACAGAGAAGATT-3' (SEQ I...

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Abstract

The invention discloses a screening or early diagnosis kit for high-risk pregnant women with gestational diabetes mellitus enormous children. The invention relates to a screening or early diagnosis kit for a high-risk pregnant woman with gestational diabetes mellitus enormous children. The kit is characterized by comprising a reagent for detecting LncRNA AC015961.2 from a serum exosome. The specificity of the LncRNA AC015961.2 derived from the serum exosome on early diagnosis of the gestational diabetic megatherium can reach 87.5%, and the sensitivity of the LncRNA AC015961.2 can reach 75.0%. According to the present invention, the expression level of the LncRNA AC015961.2 in the serum exosome of the pregnant woman suffering from gestational diabetes can be detected, and the kit can be used for the auxiliary early diagnosis of the gestational diabetes mellitus megatherium, such that the support can be provided for the clinical doctor to accurately predict the occurrence risk of the megatherium and timely adopt the more individualized prevention and treatment scheme so as to furthest reduce the adverse gestational outcome.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and relates to a screening or early diagnosis kit for pregnant women with high risk of gestational diabetes macrosomia. Background technique [0002] Macrosomia refers to infants whose birth weight reaches or exceeds 4000 grams and is the most common adverse pregnancy outcome of gestational diabetes mellitus (GDM). The latest statistics show that more than 15% of live births worldwide are affected by high blood sugar during pregnancy. In recent years, with the improvement of living standards in my country, more and more pregnant women are overweight or elderly, and the incidence of GDM is increasing year by year. GDM macrosomia is mostly of metabolic asymmetry type, which is manifested by a significant increase in the amount of body fat, while the volume of other tissues / organs is not significantly increased, which not only affects the growth and development of infants, but also gre...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/6851C12N15/11
CPCC12Q1/6883C12Q1/6851C12Q2600/178C12Q2600/158C12Q2531/113C12Q2545/101C12Q2563/107
Inventor 文娟石中华李景云马洁桦李欣邰雯
Owner NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL
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