Construction method for stably expressing Zika virus replicon containing renilla luciferase Rluc and application thereof

A technology of Renilla luciferase and Zika virus, applied in the field of Zika virus replicon construction, can solve problems such as urgent research

Pending Publication Date: 2020-06-16
LIAONING UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there are currently no effective antiviral drugs and vaccines that can be used for the clinical treatment of ZIKV, so research on safe and effective ZIKV antiviral drugs and vaccines is urgent

Method used

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  • Construction method for stably expressing Zika virus replicon containing renilla luciferase Rluc and application thereof
  • Construction method for stably expressing Zika virus replicon containing renilla luciferase Rluc and application thereof
  • Construction method for stably expressing Zika virus replicon containing renilla luciferase Rluc and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050](1) Construction method for stably expressing Zika virus replicon (Rluc-ZIKV-Rep) containing Renilla luciferase

[0051] Build methods such as figure 1 shown.

[0052] Zika virus ZIKV infectious cloning plasmid adopts ZIKVwt-FL preserved in our laboratory.

[0053] 1. Cloning of CMV-5'UTR-C22 fragment

[0054] Primer 1:

[0055] 5'-GGCGGATCCCATTGATTATTGACTAGTTATTAATAGTAA-3'

[0056] Primer 2:

[0057] 5'-CTTTCGAAGTCATGGCTACTCCGCGTTTTAG-3'

[0058] Using the plasmid ZIKVwt-FL as a template, amplify the Zika virus CMV-5'UTR-C22 fragment (22 amino acids at the 5' end of the C protein corresponding to the gene sequence), specifically:

[0059] The plasmid ZIKVwt-FL was used as a template, primer 1 and primer 2 were used as specific primers, and the Kpn I restriction enzyme site was introduced through primer design, and PCR amplification was performed to obtain the Zika virus CMV-5'UTR-C22 fragment.

[0060]

[0061] The PCR reaction conditions were as follows: 95°C...

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Abstract

The invention discloses a construction method for stably expressing a Zika virus replicon containing renilla luciferase Rluc and an application thereof. According to the invention, a replication subsystem of the Zika virus containing the renilla luciferase Rluc is constructed, so that the replication subsystem can be stably expressed in cells, and the replication condition of the virus can be detected by detecting a renilla luciferase Rluc signal. Meanwhile, the method can be used for antiviral drug screening.

Description

technical field [0001] The invention relates to the construction of a Zika virus (ZIKV) replicon, in particular to the construction and application of a Zika virus ZIKV replicon stably expressing Renilla luciferase Rluc. Background technique [0002] Zika virus (ZIKV) is an arbovirus-borne virus. It is a single-stranded positive-sense RNA virus with dengue virus, yellow fever virus, and West Nile virus. It belongs to the genus Flaviviridae and the family Flaviviridae. Since it was isolated from the serum of febrile sentinel rhesus monkeys in April 1947, it has caused outbreaks and spread in many countries and regions, and is associated with severe clinical manifestations and congenital malformations. However, there are currently no effective antiviral drugs and vaccines available for clinical treatment of ZIKV, so research on safe and effective ZIKV antiviral drugs and vaccines is urgent. [0003] Zika virus is a single-stranded positive-sense RNA virus with a length of abo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/53C12Q1/70C12Q1/02C12R1/93
CPCC12N7/00C12N9/0069C12Q1/025C12Y113/12005C12N2770/24151C12N2770/24121G01N2333/185
Inventor 刘娜胡媛媛陈凯丽郑方亮
Owner LIAONING UNIVERSITY
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