siRNA of schistosoma japonicum katsurada SjFrzb2 gene and application thereof

A technology for schistosomiasis and schistosomiasis, applied in the fields of molecular biology and biomedicine, can solve the problems of non-specific and comprehensive gene expression inhibition and apoptosis of cells

Active Publication Date: 2019-12-13
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

Moreover, dsRNA larger than 30 bp cannot induce specific RNA interference in mammals, but instead induces non-specific and comprehensive gene expression suppression and apoptosis in cells

Method used

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  • siRNA of schistosoma japonicum katsurada SjFrzb2 gene and application thereof
  • siRNA of schistosoma japonicum katsurada SjFrzb2 gene and application thereof
  • siRNA of schistosoma japonicum katsurada SjFrzb2 gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1: In vivo screening of small RNAs

[0019] 1. Method steps

[0020] 1.1 siRNA molecules and SjFrzb2 Preparation of primers for quantitative real-time PCR of genes

[0021] Find the Schistosoma Frzb2 gene sequence (accession number AAX24900.2) on the NCBI website (http: / / www.ncbi.nlm.nih.gov / ), design and synthesize three pairs of dsRNA, the design principles mainly include the following aspects: 1. 1. Starting from the AUG start codon of the transcript (mRNA), look for the "AA or NA" double-linked sequence, and write down the 19 bases at the 3' end to design; 2. Avoid starting codons or nonsense regions Select the target sequence nearby; 3. The GC content of the sequence should be about 30%-60%; 4. Compare the selected sequence in the public database to ensure that the target sequence has no homology with other genes. Simultaneously synthesize a pair of unrelated control dsRNA (NC siRNA).

[0022] S1 dsRNA: sense: 5'-GCGCACAGAUUGCAUAUAUTT-3'(SEQ ID NO.1) ...

Embodiment 2

[0041] Embodiment 2: RNA interference experiment

[0042] 1. Method steps

[0043] 1.1 dsRNA injection

[0044] The experiment consisted of control group, unrelated control group (NC group) and S1 dsRNA treatment group, with three mice in each group. 4 days, 8 days, 12 days, 16 days, 20 days, 24 days, 28 days, 32 days, 36 days and 40 days after infection, 200 µl PBS, 200 µl NC dsRNA (8 µg) DEPC water and 200 µl SjFrzb2 Gene S1 dsRNA (8 µg) DEPC water, 10 times in total.

[0045] 1.2 Liver eggs and miracidia count

[0046] Collect the liver and weigh it, add dechlorinated water, mix with a homogenizer, and set the volume to 20 mL, take 1 mL of liver homogenate, add an equal volume of 10% NaOH (W / V), and digest in a 56°C water bath After 2 h, mix well after digestion is complete, take three 50 uL samples, and count eggs under microscope. Take 4 mL of liver homogenate in a narrow-necked flat-bottomed flask, add dechlorinated water, and cover it with a thin layer of cotton at...

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Abstract

The invention discloses siRNA of a schistosoma japonicum katsurada SjFrzb2 gene. The siRNA has nucleotide sequences as shown in SEQ ID NO. 1 and SEQ ID NO. 2. The invention also discloses an application of the siRNA of the schistosoma japonicum SjFrzb2 gene. The siRNA of a schistosoma japonicum SjFrzb2 gene can obviously inhibit the transcription of the SjFrzb2 gene, the insect body number of schistosomiasis infected mice, the number of liver eggs, the liver egg contribution rate and the egg hatching rate of each female insect can be remarkably reduced, a certain pathological influence is alsocaused to the integument structure of the insect and the cell morphology of the gonad, and the product is suitable for being prepared into drugs for preventing and treating schistosomiasis.

Description

technical field [0001] The invention relates to the technical fields of molecular biology and biomedicine, in particular to a Schistosoma japonicum SjFrzb2 Gene siRNA and its application. Background technique [0002] Schistosomiasis is a zoonotic parasitic disease caused by schistosome infection. It is one of the most serious parasitic diseases in the world. More than 200 million people are infected, and nearly 800 million people are at risk of infection. Caused thousands of deaths. At present, the main prevention and treatment drug for this disease is praziquantel, but this drug cannot prevent re-infection, and long-term use has the risk of developing drug resistance. Therefore, it is imminent to prepare new vaccines or drugs for treating schistosomiasis. [0003] Frzb2 (frizzled motif associated with bone development 2) is a member of the sFrp (secret frizzled related protein) family. This gene was cloned for the first time in Xenopus laevis, and its expression was als...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113A61K31/713A61P33/12
CPCC12N15/113A61K31/713A61P33/12C12N2310/14Y02A50/30
Inventor 金亚美程贵凤李肖纯秦芳林何川川刘金明李浩古少鹏
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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