Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Method for purifying plasmids

A plasmid, calcium chloride solution technology, applied in DNA preparation, recombinant DNA technology and other directions, can solve the problems of ineffective removal of pollutants, increase removal costs, etc., and achieve the effects of removing RNA pollution, simple operation, and cost reduction

Pending Publication Date: 2022-03-11
NANJING BIOHENG BIOTECH CO LTD
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, some methods commonly used at present cannot effectively remove these pollutants, especially RNA pollutants, or need to add reagents such as enzymes and detergents, which increases the cost of removal

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for purifying plasmids
  • Method for purifying plasmids

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Embodiment 1. Purify plasmid with the method of the present invention

[0018] 1. Alkaline lysis of E. coli culture: resuspend the E. coli culture containing the plasmid to be purified evenly with resuspension buffer (TE buffer, pH8.0), and then add an equal proportion of lysate (0.2M NaOH+2 % SDS solution), after 5 minutes, add an equal proportion of 3M potassium acetate buffer to neutralize to obtain 3L lysate;

[0019] 2. Divide the lysate into 5 parts, 600ml each, add 125ml of ultrapure water to one of them (that is, the concentration of calcium chloride is 0, sample 1), and add 125ml of various concentrations of CaCl 2 Solution (pH5.0-7.0), so that the final calcium chloride concentration in the system is 0.1M (sample 2), 0.3M (sample 3), 0.5M (sample 4), 1M (sample 5).

[0020] 3. After standing at room temperature for 30 minutes, take 10 μl of the supernatant for electrophoresis (for electrophoresis results, see figure 1 1-5 in ), and take 400ml supernatant, c...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention relates to a plasmid purification method for industrial production, which comprises the following steps: (1) splitting a bacterial culture containing plasmids to be purified to obtain a lysate; (2) adding a calcium chloride solution with the final concentration of 0.5-1M into the lysate to obtain a crude extract; (3) filtering the crude extract by using a membrane filter to obtain filtrate; and (4) carrying out chromatography on the filtrate by using an anion exchange column to obtain purified plasmids.

Description

technical field [0001] The present invention describes a method for purifying plasmids. This method is mainly used for plasmid preparation in pilot scale and industrial production in the field of biopharmaceuticals. Background technique [0002] Plasmid is an extrachromosomal genetic element widely present in nature. As a carrier of genetic information, plasmids have been widely used in gene transfer operations. Especially in recent years, with the rapid development of advanced therapeutic methods (including gene therapy and cell therapy), the production of plasmids has gradually shifted from laboratory to industrial mass production. [0003] In the industrialized large-scale production of plasmids, many challenges are faced, including improving the purity of plasmids and obtaining pure plasmids that meet pharmaceutical requirements. According to the requirements of current regulations, the final product of plasmids should remove as much pollutants as possible, such as ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N15/10
CPCC12N15/1017C12N15/101
Inventor 李彦林陈明曦任江涛贺小宏王延宾韩露
Owner NANJING BIOHENG BIOTECH CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com