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Gestational diabetes mellitus micro RNA marker combination and application thereof

A technology of gestational diabetes and markers, applied in the field of biomedicine, can solve the problems of prone to false positives, slow gene mutation, and decreased detection accuracy of markers, and achieve the effect of improving detection sensitivity

Active Publication Date: 2020-01-03
SOUTHWEST MEDICAL UNIVERISTY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, due to the insufficient detection accuracy of a single microRNA, false positive or false negative results are prone to occur. At present, most of the gestational diabetes is a combination of different microRNAs, such as miR-222 , miR-132, miR-29a combination, miR-375 and miR-29 combination, but no combination of miR-132 and miR-375
Although the currently disclosed compositions have a certain potential for diagnosing gestational diabetes, due to the gradual occurrence of gene mutations, some detection markers will be eliminated or the detection accuracy will decrease, so the combination of miR-132 and miR-375 was developed It is particularly necessary as a new diagnostic marker for gestational diabetes, and its application in the preparation of diagnostic kits for gestational diabetes will have important clinical significance

Method used

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  • Gestational diabetes mellitus micro RNA marker combination and application thereof
  • Gestational diabetes mellitus micro RNA marker combination and application thereof
  • Gestational diabetes mellitus micro RNA marker combination and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Detection of miR-132 and miR-375 in the peripheral blood of pregnant women with gestational diabetes

[0027] For miR-132 and miR-375 detection, extract serum RNA according to the instructions of the RNA extraction kit (Invitrogen Ambion), perform reverse transcription according to the instructions of the TaqMan reverse transcription kit (APPS, USA), and then use the reverse transcription product as a template. Perform RT-PCR reactions on different samples.

[0028] The upstream primer for amplifying miR-132 is: 5'-ACACTCCAGCTGGGTAGCAGCACGTAAATA-3', shown in SEQ ID NO. 1,

[0029] The downstream primer is: 5'-CTCAACTGGTGTCGTGGAGTCGGCAATTCAGTTGAGCGCCAATA-3', SEQ ID NO. 2;

[0030] The upstream primer for amplifying miR-375 is: 5'-CCCTCTAGACCCCAAGGCTGATGCTGAGAAG-3', SEQ ID NO. 3, and the downstream primer 5'-AAAGGTCCGCCGCCCGGCCCCGGGTCTTC-3', SEQ ID NO. 4.

[0031] The RT-PCR amplification conditions of miR-132 and miR-375 are the same, 95°C for 10 min; 95°C for 15s, 95°C...

Embodiment 2

[0036] Example 2 The effect of different RT-PCR reaction systems on the relative expression of miR-132 and miR-375 in non-gestational diabetes patients.

[0037] Using the cDNA obtained by reverse transcription of RNA extracted from the peripheral blood serum of non-gestational diabetes patients obtained in Example 1 as a template, different RT-PCR reaction systems were used for detection, miR-132, miR-375 amplification primers and The amplification conditions were the same as in Example 1. The different reaction system settings are as follows:

[0038] Reagent blank group: use the 2×SYBR Green Mix reaction kit of Bori Technology Company, and perform RT-PCR of 21μL system according to the instructions of the kit. DdH for cDNA in reagent blank group 2 O dissolves, the final concentration of cDNA is 20ng / μL.

[0039] Reagent experimental group: the reaction system is basically the same as the reagent blank group, the difference is that the ddH is replaced with an equal volume of sodi...

Embodiment 3

[0044] Example 3 The effect of different RT-PCR reaction systems on the relative expression of miR-132 and miR-375 in patients with gestational diabetes.

[0045] In Example 3,

[0046] In Example 3, cDNA obtained by reverse transcription of RNA extracted from the peripheral blood serum of a gestational diabetes patient obtained in Example 1 was used as a template, and different RT-PCR reaction systems were used for detection. The rest of the test conditions were the same as in Example 2. The results are shown in Table 4. In Table 4, in the reaction system of the sodium chloride-potassium chloride complex reagent, the relative expression levels of miR-132 and miR-375 are increased, indicating that the reaction actually has a higher response value and is better suited for blood The low content of miR-132 and miR-375 and the small amount of extracted cDNA template increase the sensitivity of the detection result.

[0047] Table 4 Relative expression of microRNA under different reacti...

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Abstract

The invention belongs to the technical field of biomedicine, and particularly discloses a micro RNA marker combination related to gestational diabetes and application thereof. The micro RNA marker combination comprises miR-132 and miR-375. The invention further discloses a preparation method of the micro RNA marker combination. The miR-132 and the miR-375 are located in peripheral blood serum or blood plasma. The miR-132 and the miR-375 can be used as diagnostic markers for the gestational diabetes, and a basis is provided for early diagnosis of the gestational diabetes mellitus.

Description

Technical field [0001] The invention belongs to the field of biomedical technology, and specifically relates to providing a combination of microRNA markers for gestational diabetes and its application. Background technique [0002] Gestational diabetes is a complication of pregnancy. The current clinical diagnosis is mainly based on the detection of pregnant women's blood glucose in the second trimester. Since gestational diabetes may cause fetal malformations, and severe cases endanger the health of mothers and infants, the earlier the diagnosis of gestational diabetes is, the better it is for the patient. . [0003] MicroRNA (microRNA) is a type of small non-coding RNA with a molecular weight of 18-25 bases. MicroRNA is highly conserved in evolution and participates in the regulation of gene expression after transcription. It regulates gene expression and its expression at the post-transcriptional and translational levels. The abnormality is closely related to gestational diabet...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C12N15/11
CPCC12Q1/6883C12Q2600/178C12Q2600/158
Inventor 罗茂方丹陈然贺朝晖吴剑波
Owner SOUTHWEST MEDICAL UNIVERISTY
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