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Non-coding small RNA molecular spectrum related to tumor lactic acid microenvironment and application thereof

A technology of small molecules and lactic acid, which is applied in the determination/testing of microorganisms, biochemical equipment and methods, etc., and can solve the problems of difficult acquisition of pathological specimens, infection, necrosis, etc.

Pending Publication Date: 2021-06-11
FUDAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In clinical practice, the clinical diagnosis and prognosis of lymphoma are mainly carried out through invasive lymph node excision biopsy combined with immunohistochemistry, etc. Since the disease site can appear in any tissue in or outside the lymph nodes of the whole body, pathological specimens are not easy to obtain; In addition, lymphoma lesions are often accompanied by infection, necrosis, etc., which makes pathological diagnosis difficult and requires multiple biopsies

Method used

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  • Non-coding small RNA molecular spectrum related to tumor lactic acid microenvironment and application thereof
  • Non-coding small RNA molecular spectrum related to tumor lactic acid microenvironment and application thereof
  • Non-coding small RNA molecular spectrum related to tumor lactic acid microenvironment and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0101] Example 1: Screening and Identification of Lactic Acid Microenvironment Responsive miRNA Molecules

[0102] Using stem-loop qRT-PCR, in EBV-positive cell lines (LCL, B95.8) and EBV-negative cell lines (BJAB), the expression levels of miRNA molecules under different concentrations of lactic acid were detected, and a group of lactic acid-responsive and A miRNA expression profile with a stable trend. This group of miRNAs is hsa-miRNA-7974, hsa-miRNA-7-5p, hsa-miR-4521, hsa-miRNA-9-5p, hsa-miRNA210-3p ( image 3 ).

[0103] Detection of miRNA-stem-loop real-time quantitative PCR: a stem-loop reverse transcription primer is designed separately for each miRNA, and the 5' end of the stem-loop RT primer specifically binds to the 3' end of the miRNA molecule. Under the action of the enzyme, the lengthened miRNA first-strand cDNA is obtained. The obtained reverse transcription product is subjected to qPCR reaction under the action of qPCR specific upstream primer and downstrea...

Embodiment 2

[0136] Example 2: Lactic acid microenvironment responds to 5'-tiRNA HisGUG screening and identification of

[0137] A 5'-tRNA moiety: 5'-tRNA was screened out in EBV-positive cell lines (LCL1 and B95.8) by fluorescent quantitative PCR HisGUG , its expression has a certain trend of change with the increase of lactic acid concentration. At the same time, the expression level of ANG molecules upstream of the cleaved tRNA was detected, and the results showed that the expression level of ANG also changed with the increase of lactic acid concentration ( Figure 7 ). This suggests that 5'-tRNA HisGUG Lactic acid plays a certain role in promoting the occurrence of B-cell lymphoma, and has potential clinical value in the diagnosis and prognosis of EBV-related B-cell lymphoma.

[0138]Detection and quantitative analysis of tRNA-half molecules: The structural feature of 5-tiRNA is that there are 2', 3'-cyclic phosphate groups at the 3' end. In the first step, T4 PNK treats the total ...

Embodiment 3

[0156] Example 3: Detection of expression levels of hsa-miR-9-5p, hsa-miR-210-3p, hsa-miR-7-5p, hsa-miR4521, hsa-miR-7974 in 8 lymphoma samples

[0157] Eight clinical samples (PBMC) of lymphoma and other diseases with EBV negative and normal LDH (lactate dehydrogenase) levels ( Figure 7 .

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Abstract

The invention belongs to the technical field of biochemistry and molecular biology, and particularly relates to amplification and analysis of hsa-miR-9-5p, hsa-miR-210-3p, hsa-miR-7-5p, hsa-miR-4521, hsa-miR-7974 and tiRNAHisGUG coded by human cells, expression conditions thereof, and applications thereof for prediction. the invention provides a method for quantifying small molecule RNA. The method comprises the following steps: (1) collecting a sample; wherein the sample is in an environment containing lactic acid, and the concentration range of the lactic acid is 100-250U / L; (2) performing quantitative PCR by a stem-loop method; and (3) obtaining a target miRNA molecule or a 5'-tiRNA molecule. The invention also provides application of the target micromolecule RNA, and provides a new tool for prognosis effect and treatment of malignant lymphoma.

Description

technical field [0001] The invention belongs to the field of biochemistry and molecular biology, and specifically relates to the amplification and analysis of hsa-miR-9-5p, hsa-miR-210-3p, hsa-miR-7-5p, hsa-miR4521, hsa encoded by human cells -miR-7974 and tiRNA HisGUG , and its expression, and its application in predicting the prognosis and treatment of malignant lymphoma. Background technique [0002] The prior art discloses that small RNAs include microRNA (miRNA), transfer RNA (tRNA), Piwi protein-associated RNA (piRNA), small nucleolar RNA (snoRNA), small nuclear RNA (snRNA) and ribosomal RNA (rRNA), etc. . miRNA is a kind of non-coding small RNA with a length of about 18-25nt. Its expression in vivo is sequential, conservative, tissue-specific and disease-specific. It can negatively regulate the expression of mRNA by complementary binding with target genes. It plays a role in various life activities such as cell proliferation and differentiation, apoptosis, embryoni...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12Q1/6851
CPCC12Q1/6851C12Q1/6886C12Q2600/118C12Q2600/158C12Q2600/178C12Q2531/113C12Q2525/207C12Q2525/301C12Q2527/125
Inventor 蔡启良陈小亭莫小辉都树娟朱彩霞王玉燕
Owner FUDAN UNIV
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