Method for producing non-enveloped viral particles

Patent metadata

Field	Value
Publication number	US-10415020-B2
Application number	US-201615540606-A
Country	US
Kind code	B2
Filing date	Jan 8, 2016
Priority date	Jan 9, 2015
Publication date	Sep 17, 2019
Grant date	Sep 17, 2019

Abstract

Provided is a method for producing non-enveloped viral particles, comprising a step for obtaining a fraction containing non-enveloped viral particles by removing precipitates which are produced in a step for adding a substance which reduces the solubility of proteins under acidic conditions and/or a substance which precipitates under acidic conditions to a neutral or basic sample containing non-enveloped viral particles, and acidifying the sample after the addition of the substance.

First claim

The invention claimed is:

1. A method of producing a non-enveloped virus particle, the method comprising:
(a) a step of adding a carboxylic acid type anionic surfactant to a neutral or basic sample containing a non-enveloped virus particle,
(b) a step of acidifying the sample to which the surfactant is added to form a precipitate,
(c) a step of removing a precipitate formed in step (b) to obtain a supernatant fraction containing the non-enveloped virus particle,

wherein the non-enveloped virus particle is adeno-associated virus particle.

2. The method according to claim 1 , which further comprises (d) a step of purifying the non-enveloped virus after step (c).

3. The method according to claim 1 , wherein the carboxylic acid type anionic surfactant is selected from the group consisting of sodium deoxycholate, sodium chenodeoxycholate, sodium cholate, sodium glycocholate, sodium taurocholate, and sodium taurodeoxycholate.

4. The method according to claim 1 , wherein the step of acidifying is a step of adding an acidic solution.

5. The method according to claim 1 , wherein the neutral or basic sample containing a non-enveloped virus particle is a lysate or a homogenate of a non-enveloped virus-producing cell, a culture supernatant of a non-enveloped virus-producing cell which contains a non-enveloped virus particle, or an extract obtained from a non-enveloped virus-producing cell.

6. The method according to claim 5 , wherein the neutral or basic sample containing a non-enveloped virus particle is a sample obtained by bringing a non-enveloped virus-producing cell into contact with an acidic solution and then adjusting to a neutral or basic pH.

7. The method according to claim 1 , wherein the neutral or basic sample containing a non-enveloped virus particle is a sample treated with nuclease.

8. A kit for performing the method according to claim 1 , comprising:
(1) a carboxylic acid type anionic surfactant, and
(2) an acidic solution.

9. The kit according to claim 8 , which further comprises a neutralizing solution.

Assignees

• Takara Bio Inc

Inventors

• Kawano Yasuhiro
• Huang Shuohao
• Enoki Tatsuji
• Kitagawa Masanari

Classifications

• C12N15/09

  Recombinant DNA-technology · CPC title

• C12N7/02

  Recovery or purification · CPC title

• C12N5/10

  Cells modified by introduction of foreign genetic material · CPC title

• C07K14/075

  Adenoviridae · CPC title

• C12N2750/14121

  Viruses as such, e.g. new isolates, mutants or their genomic sequences · CPC title