"Low-Salt Lysis Buffer" Preparation
Updated 6/12/2015 08:34 PM
Introduction
Materials
- Disposables
- weigh dishes/paper
- pipette tips
- Equipment
- sterile container
- balance
- pipette
Procedure
Components
Table1
A A | B B | |
1 1 | Final Volume (ml)* | 100 |
Table2
A A | B B | C C | D D | |
1 1 | Component | Stock Conc. (M)* | Amount | Unit |
2 2 | Tris-Cl (pH 8.5) | 1.0 | 1.000 | ml |
3 3 | EDTA (pH 8.0) | 0.5 | 0.400 | ml |
4 4 | lysozyme (optional) | 10 | mg | |
5 5 | dH2O | to final volume |
Preparation
- Add ~70% of the final volume of sterile dH2O to a sterile container.
- Add calculated amount of each component in the tables above in order.
- If using lysozyme, use immediately; otherwise, store at room temperature for short term.
Non-lysozyme solution can be autoclaved if desired.
This protocol describes the preparation of the lysis buffer used in the dual ELP-tagged split-intein system of protein expression and purification.
This buffer may possess lysozyme, or it may be omitted as in Shi et al. 2013. If using lysozyme, prepare immediately before use.
Fong, B. A., Wu, W.-Y. & Wood, D. W. (2009). Optimization of ELP-intein mediated protein purification by salt substitution. Protein Expr Purif 66, 198-202.
Shi, C., Meng, Q. & Wood, D. (2013). A dual ELP-tagged split intein system for non-chromatographic recombinant protein purification. Appl Microbiol Biotechnol 97, 829-835.