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Hsa-let-7 familial miRNA and applications of target genes of hsa-let-7 familial miRNA in diagnosis and treatment for EBOV infection

A technology of hsa-let-7, target gene, applied in the direction of disease diagnosis, biochemical equipment and methods, microbial determination/inspection, etc.

Inactive Publication Date: 2017-03-29
MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, many studies on virus pathogenicity are carried out in animal models and in vitro experiments, but there are few related studies on human virus infection

Method used

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  • Hsa-let-7 familial miRNA and applications of target genes of hsa-let-7 familial miRNA in diagnosis and treatment for EBOV infection
  • Hsa-let-7 familial miRNA and applications of target genes of hsa-let-7 familial miRNA in diagnosis and treatment for EBOV infection
  • Hsa-let-7 familial miRNA and applications of target genes of hsa-let-7 familial miRNA in diagnosis and treatment for EBOV infection

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Experimental program
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Effect test

Embodiment 1

[0059] Example 1. Host key molecules involved in the regulation of EBOV-infected patient cells

[0060] Extract total RNA from the whole blood of EBOV-infected patients and non-EBOV-infected patients, use magnetic beads with Oligo(dT) to absorb and purify the mRNA in the total RNA, fragment the mRNA under heating conditions, and use this as a template for inversion Recorded into double-stranded cDNA. The double-stranded cDNA is repaired and filled to phosphorylate the 5' end and add "A" to the 3' end. The double-stranded cDNA adapter with 3'dTMP end was connected to the sequencing adapter, amplified by PCR, enriched and purified with AMPure XP magnetic beads, and the library was identified by PCR reaction. The constructed library was subjected to paired-end sequencing using the Illumina sequencing platform. Use the Perl script to filter adapter sequences, low-quality sequences at both ends (sequences with Q<=20 bases accounting for more than 50% of the entire reads), and low...

Embodiment 2

[0072] Embodiment 2, small RNAs (sRNAs) involved in EBOV infection damage regulation

[0073] Extract total RNA from the whole blood of EBOV-infected patients and non-EBOV-infected patients, use 15% denatured polyacrylamide gel electrophoresis (PAGE) to separate small fragments (15-30nt) RNA from total RNA, and isolate small fragments after purification The fragmented RNA was ligated with adapters at the 3' and 5' ends, and then the RNA with the adapters was reverse-transcribed using SuperScriptII reverse transcriptase (product of Invitrogen) to synthesize cDNA. Then, PCR amplification was carried out, and the amplification program was: pre-denaturation at 98°C for 30s, denaturation at 98°C for 10s, annealing at 60°C for 30s, extension at 72°C for 15s, 14 cycles, and extension at 72°C for 8min, thereby constructing the library. Finally, the extended products were subjected to high-throughput sequencing using the Illumina sequencing platform. Use the Perl script to filter the ...

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Abstract

The invention discloses hsa-let-7 familial miRNA and applications of target genes of the hsa-let-7 familial miRNA in diagnosis and treatment for EBOV infection. The hsa-let-7 familial miRNA disclosed by the invention is hsa-let-7a-5p, hsa-let-7b-5p, hsa-let-7c-5p, hsa-let-7d-5p and hsa-let-7f-5p, the target genes of the hsa-let-7 familial miRNA are MAB21L3 gene, MEIS3 gene, DPP6 gene, IGDCC3 gene, CHRD gene, FASLG gene, LIN28 gene, E2F5 gene, IL-6 gene and E2F6 gene. The hsa-let-7 familial miRNA and the target genes of the hsa-let-7 familial miRNA can be taken as biological molecular markers for diagnosing EBOV infection, and also can be taken as biological target molecules for the design of the medicines capable of resisting EBOV infection.

Description

technical field [0001] The invention relates to the application of hsa-let-7 family miRNA and its target gene in the diagnosis and treatment of EBOV infection in the field of biotechnology. Background technique [0002] Ebola virus disease is an acute hemorrhagic infectious disease caused by Ebola virus (EBOV) belonging to the filoviridae family. The fatality rate is very high, up to 50% to 90%. [0003] The Ebola virus genome is a single-stranded negative-strand RNA, about 19kb in length, which can encode seven structures including nucleoprotein (NP), envelope protein (VP35, VP40, VP30, VP24), glycoprotein (GP) and RNA polymerase Protein, among which GP gene has unique coding and transcriptional functions for EBOV replication. The virus replicates, assembles, and releases by budding in the cytoplasm of infected cells. Ebola virus mainly includes Zaire type (EBOV-Z), Sudan type (EBOV-S), Côte d'Ivoire type (EBOV-C) and Reston type (EBOV-R), etc. Different subtypes have di...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N33/68
CPCC12Q1/6883C12Q2600/158C12Q2600/178G01N33/6893G01N2800/26
Inventor 王慧李涛刘雄刘坤宁年智
Owner MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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