Method of purifying Bt toxalbumin from insect-resistant cotton seeds
A Bt poisonous protein and insect-resistant cotton technology, applied in the field of biochemistry, can solve the problems reflecting the harm of Bt gene crops, changes in protein properties and functions, etc.
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
preparation example Construction
[0065] Preparation method of cell culture medium: add calf serum (purchased from GIBCOBRL, catalog number 26170-043) and sodium bicarbonate to DMEM medium, the final concentration of calf serum is 20% (mass percentage content), The final concentration of sodium bicarbonate was 0.2% (mass percentage), and the pH was 7.4.
[0066] Bt Cry1Ac protein: Shanghai Youlong Biotechnology Co., Ltd., catalog number C010301.
[0067] Cotton variety Shiyuan 321: Hebei Shennong High-tech Co., Ltd.
[0068] The seeds of Guoxin Mian No. 6 (an insect-resistant variety transfected with the gene encoding the Bt Cry1Ac protein) were purchased from the Guoxin Agricultural Research Association in Hejian City, Hebei Province.
[0069] Unless otherwise specified, the PBS buffer used in this embodiment is as follows: the formula of PBS buffer (pH7.5): the solvent is water, containing 0.02M Na 2 HPO 4 、0.0015M KH 2 PO 4 and 0.14M NaCl.
Embodiment 1
[0070] Embodiment 1, purify Bt Cry1Ac protein from insect-resistant cotton seeds
[0071] 1. Extraction of total soluble protein
[0072] The seeds of Guoxin Cotton No. 6 were dehulled and ground into a fine powder. Take 10 g of the fine powder, add 100 ml of pH10.5, 0.01 MCAPS buffer, stir and extract with a magnetic stirrer at 4°C for 4 hours, and then centrifuge at 4°C and 10,000rpm for 20 Minutes, avoid the surface oil and take the upper liquid, which is the total soluble protein extract.
[0073] 2. Ammonium sulfate precipitation
[0074] Add ammonium sulfate powder to the soluble total protein extract obtained in step 1 to make its saturation in the extract reach 60%, and stir for 40 minutes in an ice bath using a magnetic stirrer.
[0075] 3. Reconstitution and dialysis of precipitate
[0076] Centrifuge the extract that completed step 2 at 4°C and 7000rpm for 20 minutes to collect the precipitate; add 2ml of deionized water to the precipitate, adjust the pH to 10.5 ...
PUM
Abstract
Description
Claims
Application Information
- R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
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