TSS: Difference between revisions
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==Uses== | |||
TSS buffer is used to make [[Preparing chemically competent cells|chemically competent cells]]. | |||
==Preparation== | |||
To make 50 mL: | To make 50 mL: | ||
*5g PEG 8000 | *5g PEG 8000 | ||
*1.5 mL 1M | *1.5 mL 1M MgCl<sub>2</sub> (or 0.30g MgCl<sub>2</sub>*6H<sub>2</sub>0) | ||
*2.5 mL DMSO | *2.5 mL DMSO | ||
*LB to 50 mL | *Add LB to 50 mL | ||
Filter sterilize (0.22 μm filter) | |||
==Notes== | |||
*Note 1: PEG 3350 can be used instead of PEG 8000. According to the original CT Chung paper, PEG 3350 produces better efficiency, and other sized PEGs can be used as well albeit with a slight loss in efficiency. | |||
*Note 2: pH from original Chung CT paper calls for pH to be acidic (pH=6.5)... be sure to pH solution before sterilization. | |||
*Note 3: If using non-chemically resistant filters (e.g., cellulose nitrate), add DMSO after sterilization. DMSO should be sterile in and of itself, so it may be prudent it add it afterwards if you are unsure about the compatibility of your filters. | |||
Store at | ==Storage== | ||
Store at 4˚C or -20˚C. | |||
[[Category:Escherichia coli]] [[Category:Material]] [[Category:Buffers]] | |||
Latest revision as of 11:08, 4 April 2013
Uses
TSS buffer is used to make chemically competent cells.
Preparation
To make 50 mL:
- 5g PEG 8000
- 1.5 mL 1M MgCl2 (or 0.30g MgCl2*6H20)
- 2.5 mL DMSO
- Add LB to 50 mL
Filter sterilize (0.22 μm filter)
Notes
- Note 1: PEG 3350 can be used instead of PEG 8000. According to the original CT Chung paper, PEG 3350 produces better efficiency, and other sized PEGs can be used as well albeit with a slight loss in efficiency.
- Note 2: pH from original Chung CT paper calls for pH to be acidic (pH=6.5)... be sure to pH solution before sterilization.
- Note 3: If using non-chemically resistant filters (e.g., cellulose nitrate), add DMSO after sterilization. DMSO should be sterile in and of itself, so it may be prudent it add it afterwards if you are unsure about the compatibility of your filters.
Storage
Store at 4˚C or -20˚C.
