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Gestational diabetes biomarkers of intestinal bacteria in early pregnancy as well as screening and application of gestational diabetes biomarkers

A technology of biomarkers and intestinal bacteria, which is applied in the fields of biomaterial analysis, biochemical equipment and methods, and microbial measurement/inspection, etc. It can solve the problems of small sample size, shortage of clinical data, unreasonable selection of control groups, etc. problem, achieve affordability guarantee and improve forecasting performance

Inactive Publication Date: 2021-07-27
HUAZHONG UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the design of this study is not focused on GDM, and the relevant data obviously have defects such as too small sample size, unreasonable selection of the control group, and shortage of epidemiological and other clinical data.

Method used

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  • Gestational diabetes biomarkers of intestinal bacteria in early pregnancy as well as screening and application of gestational diabetes biomarkers
  • Gestational diabetes biomarkers of intestinal bacteria in early pregnancy as well as screening and application of gestational diabetes biomarkers
  • Gestational diabetes biomarkers of intestinal bacteria in early pregnancy as well as screening and application of gestational diabetes biomarkers

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Example 1: Isolation and purification of stool sample DNA

[0058] (1) Configuration of bacterial lysate

[0059] Before extracting fecal DNA, the bacterial lysate is prepared to break up the cells in the fecal tissue, so that the internal bacterial DNA can be released. Prepare the bacterial lysate in a 50mL centrifuge tube according to the following system.

[0060] Bacterial lysate system (32mL):

[0061]

[0062] After vortex mixing, dispense into 8 ultrafiltration tubes, 4 mL in each tube. After centrifugation at 3500 rpm / min at 4°C for 30 min, the filter membrane was discarded to obtain a bacterial lysate. Transfer to four 15mL centrifuge tubes and store at -20°C.

[0063] (2) Fecal DNA extraction

[0064] Before DNA extraction, fecal samples were centrifuged at 12,000 rpm / min for 10 min to remove ethanol, and then the Tiangen Fecal DNA Extraction Kit (DP328) was used to extract fecal DNA. Weigh 0.18-0.22g of stool sample into a 2mL Safe-lock centrifuge tu...

Embodiment 2

[0071] Example 2: 16S rRNA gene amplicon library construction

[0072] The V3-V4 region of the 16S rRNA gene was amplified by two-step PCR to construct a library.

[0073] The first step of PCR is:

[0074] The primer sequence is as follows: Forward (341F):

[0075] 5'-ACACTCTTTCCCTACACGACGCTCTTCCGATCTCCTACGGGNGGCWGCAG-3';

[0076] Reverse (805R):

[0077] 5'-AGACGTGTGCTCTTCCGATCTGACTACHVGGGTATCTAATCC-3'.

[0078] The first step of PCR amplification reaction was carried out according to the following system.

[0079] The first step PCR amplification system (25μL):

[0080]

[0081] The first step PCR reaction conditions:

[0082]

[0083] The first step PCR amplification product was purified by magnetic bead method to remove impurities such as primer dimers.

[0084] The second step of PCR is:

[0085] Primer sequence: Forward: 5'-AATGATACGGCGACCACCGAGATCTACAC (Index sequence 1)ACACTCTTTCCCTACACGACG-3';

[0086] Reverse 5'-CAAGCAGAAGACGGCATACGAGAT (Index sequenc...

Embodiment 3

[0096] The intestinal bacteria and bacterial indicators associated with GDM were screened by bioinformatics methods and statistical methods. The analysis process is briefly described as follows:

[0097] (1) Sequence data processing

[0098] The constructed library was sequenced on the Illumina Miseq platform to obtain the original sequence data, mainly through QIIME2 (Quantitative Insights into Microbial Ecology version 2, QIIME2) ( https: / / qiime2.org / ) for processing, significance analysis, and R package for visualization.

[0099] The off-machine data contains sequencing joints and some low-quality reads (Reads), and a plug-in based on QIIME2 software is required to perform quality control processing on the original off-machine data. Filtering of low-quality sequences, splicing of double-ended sequences, and removal of chimeras are all done by the DADA2 plugin of QIIME2. DADA2 is a model-based correction algorithm. Different from the traditional UPARSE algorithm, whi...

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Abstract

The invention discloses gestational diabetes biomarkers of intestinal bacteria in early pregnancy as well as screening and application of the gestational diabetes biomarkers, and belongs to the field of intestinal flora and human health research. 27 intestinal bacterium biomarkers and 4 intestinal bacterium calculation indexes are included. The sample DNA is subjected to 16S rDNA amplicon library building and sequencing, and the structure and function information of intestinal flora and the relevance with gestational diabetes are analyzed. Regression model inspection shows that the screened 6 types of intestinal bacteria significantly improve the prediction efficiency of a gestational diabetes basic prediction model. The relative abundance of the staphylococcus and the 4 types of short-chain fatty acid producing bacteria is higher than that of a log converted value in a case group, and is positively correlated with fasting blood-glucose, 1h blood-glucose level and 2h blood-glucose level in the middle pregnancy period. The biomarkers provided by the invention can be used as a detection target for preparing a kit for predicting the onset risk of gestational diabetes at the early pregnancy stage, or can be used as a target for preparing a medicine for treating and / or preventing gestational diabetes.

Description

technical field [0001] The present invention belongs to the field of intestinal flora and human health research, and more specifically, relates to a series of intestinal bacteria and bacterial calculation indicators associated with the onset of gestational diabetes in the first trimester, especially the gestational diabetes biological index of intestinal bacteria in the first trimester. Markers and screening and application. Background technique [0002] With China's rapid economic development and social transformation, maternal and child health will face new problems. Risk factors such as high-nutrient diet, low physical activity, obesity, and advanced maternal age will affect maternal health and pregnancy outcomes. Among them, Gestational Diabetes Mellitus (GDM) is one of the main complications during pregnancy, which seriously endangers the health of mother and child. Studying the pathogenesis of GDM and finding the risk factors that can be used to predict GDM early are ...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/6895C12Q1/06C12Q1/14G16H50/30G06K9/62
CPCC12Q1/689C12Q1/6895C12Q1/06C12Q1/14G16H50/30G01N2800/042G01N2800/50G06F18/241
Inventor 胡平潘安潘雄飞谌秀仪褚旭峰
Owner HUAZHONG UNIV OF SCI & TECH
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