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Micro rna and its application in the preparation of preparations against Schistosoma japonicum infection

A technology of schistosomiasis and preparations, which is applied in the field of microRNA and its preparation of anti-schistosomiasis infection preparations, which can solve problems such as difficult to control repeated infection of schistosomiasis

Active Publication Date: 2022-01-18
STATION OF VIRUS PREVENTION & CONTROL CHINA DISEASES PREVENTION & CONTROL CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, praziquantel is still the main treatment for schistosomiasis, but it is difficult to control the superinfection of schistosomiasis completely relying on single drug therapy, and in recent years, studies have found that praziquantel chemotherapy has the potential risk of inducing drug resistance, so new schistosomiasis vaccines and The development of new drugs has important strategic significance for the effective control and even elimination of schistosomiasis

Method used

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  • Micro rna and its application in the preparation of preparations against Schistosoma japonicum infection
  • Micro rna and its application in the preparation of preparations against Schistosoma japonicum infection
  • Micro rna and its application in the preparation of preparations against Schistosoma japonicum infection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0131] Example 1 Design of Schistosoma amiRNA

[0132] By screening a large number of genes, it is found that SjCL gene and FTL gene may play an important role in physiological processes such as the growth and development of Schistosoma japonicum, so the present invention is aimed at Schistosoma japonicum SjCL gene (accession number: FN319073. No.: FN318171.1) each designed four amiRNAs, and the four amiRNAs of the SjCL gene were named amiSJCL1, amiSJCL2, amiSJCL3, and amiSJCL4, respectively, and amiSJCL1, amiSJCL2, amiSJCL3, and amiSJCL4 were located in the 123bp, 283bp, 455bp, and 674bp segments of the SJCL gene, respectively. The four amiRNAs of the FTL gene were named amiFTL1, amiFTL2, amiFTL3, and amiFTL4, respectively, and amiFTL1, amiFTL2, amiFTL3, and amiFTL4 were located in the 211bp, 276bp, 354bp, and 454bp segments of the FTL gene, respectively. AmiRNAs are formed by annealing of two single-strands, the top strand and the bottom strand, to form a double strand.

[...

Embodiment 2

[0143] Example 2 Construction of recombinant lentiviral vector pSIF-H1 corresponding to amiRNA of Schistosoma japonicum ftl and sjcl genes

[0144] Because only one fragment is designed, the recombinant vector may not work, so amiFTL4, amiSJCL2, and amiSJCL4 each design two fragments with different lengths. The short fragment FTL4-miR of the two fragments of amiFTL4 only contains a short fragment of amiRNA, and the long fragment contains an additional trans element named FTL4-trans in addition to amiRNA, where trans means that it also contains an upstream transcription initiation element (in order to drive Transcription), miR represents only amiRNA short fragments, similarly the short fragment of amiSJCL2 is SJCL2-miR, the long fragment is SJCL2-trans, the short SJCL4 fragment is SJCL4-miR, the long fragment is SJCL4-trans, FTL4 and SJCL2, SJCL4 long and short fragments The upstream contains the BamHI restriction site, and the downstream contains the EcoRI restriction site. A...

Embodiment 3

[0146] Example 3 Screening of effective pSIF-recombinant vector

[0147] 3.1 Method

[0148] 293T cell culture: The previously constructed SJCL gene overexpression vector pCMV6-SJCL (purchased from Origene) was co-transfected with pSIF-SjCL4-miR and pSIF-SjCL4-trans into 293T cells, respectively, and cultured for 48 hours and 72 hours; previously constructed 293T cells were co-transfected with the FTL gene overexpression vector pCMV-FTL (purchased from Origene), pSIF-SJCL4-miR and pSIF-SJCL4-trans, and cultured for 48 hours and 72 hours, respectively.

[0149] 3.2 Transcript level and protein level identification

[0150] 3.2.1 Results of real-time fluorescent quantitative PCR (qPCR)

[0151] qPCR was used to verify the selection of fragments with better effects, and the Graphad-prism5 software was used to count the PCR results, and it was found that pSIF-SJCL4-miR and pSIF-SJCL4-trans could effectively interfere with the expression of pCMV-SjCL, as shown in figure 2 A, P ...

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Abstract

The invention provides microRNA and its application in preparation of anti-Schistosoma japonicum infection preparation. Specifically, the present invention provides nucleic acid inhibitory molecules, especially microRNA nucleic acid molecules, which have significant killing or inhibiting effects on Schistosoma japonicum. Experiments show that the specific amiRNA of the present invention has a high ability to kill or inhibit Schistosoma japonicum (especially early stage schistosomiasis). The invention also provides the preparation method, pharmaceutical composition and application of the nucleic acid inhibitory molecule.

Description

technical field [0001] The invention relates to biomedicine, in particular to microRNA and its application in the preparation of anti-schistosomiasis infection preparations. Background technique [0002] Schistosomiasis is a parasitic disease that seriously endangers human health. At present, praziquantel is still the main treatment for schistosomiasis, but it is difficult to control the superinfection of schistosomiasis completely relying on single drug therapy, and in recent years, studies have found that praziquantel chemotherapy has the potential risk of inducing drug resistance, so new schistosomiasis vaccines and The development of new drugs has important strategic significance for the effective control and even elimination of schistosomiasis. Non-coding small RNAs widely exist in eukaryotes, generally 18-30 nucleotides (nucleotides, nt) in length, mainly including microRNAs (miRNAs), small interfering RNAs (siRNAs) ), Piwi-interacting ribonucleic acid (piRNA) and ot...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N15/11A61K31/713A61K31/7088A61P33/12G01N33/68
CPCG01N33/68C12N15/1137C12N15/1138C12N2310/141Y02A50/30
Inventor 秦志强许静杨杰吕山李石柱周晓农
Owner STATION OF VIRUS PREVENTION & CONTROL CHINA DISEASES PREVENTION & CONTROL CENT
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