TSS

From OpenWetWare

(Difference between revisions)
Jump to: navigation, search
Current revision (14:08, 4 April 2013) (view source)
m (Storage)
 
(8 intermediate revisions not shown.)
Line 1: Line 1:
 +
==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
Line 5: Line 10:
*Add LB to 50 mL
*Add LB to 50 mL
-
Filter sterilize (0.22 μm filter) and store at 4˚C or -20˚C.
+
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.
 +
[[Category:Escherichia coli]] [[Category:Material]] [[Category:Buffers]]

Current revision

Contents

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.

Personal tools