Revision as of 13:29, 28 August 2013 by Batou (Talk | contribs)

(diff) ← Older revision | Latest revision (diff) | Newer revision → (diff)

Siderophore detection

Aim

Once our bacteria is transformed with the plasmid with the Fur Binding Site and Lac I and the two plasmids with Enterobactins genes, we need to check if our system really work. The production of siderophores can be detected visualy with Blue Agar Chrome Azurol S (CAS). Without siderophore in the medium, CAS and Hexadecyltrimethylammonium bromide (HDTMA)complexes with ferric iron, producing a blue color. When a bacteria strain produce siderophore, the medium color change from blue to orange.

Preparation

Protocol adapted from Louden, Haarmann, and Lynne, “Use of Blue Agar CAS Assay for Siderophore Detection.”

Blue Dye

Solution 1
Dissolve 0,06 g of CAS in 50 mL of distilled water.

Solution 2
Dissolve 0,0027 g of FeCl3-6H20 in 10 mL of 10 mM HCl.

Solution 3
Dissolve 0,073 g of HDTMA in 40 mL of distilled water.

Mix
Mix solution 1 with 9 mL of solution 2. Then mix with solution 3.
Solution should have a blue color.
Autoclave and store in a bottle.

Blue Dye

Site developped by Gabriel Guillocheau