is there a rapid method of extraction and revelation of AAI products by microbial strains ?
Hi
You can use spectrophometric method as following:
prepare Salkowski reagent (FeCl3 10mM /perchloric acid 35%)
- Centrifugation of bacterial culture suspension at 10000g for 10 min at 4°C
- Take 1 ml of free bacteria supernatant
- add 2 drop of orthophosphoric acid
- add 2 ml of Salkowski reagent
- Incubation at room T° for 20-25 min
in presence of IAA, we see change the color to pink
Take OD at 535 nm against a blank prepared in the same condition with bacterian medium
you can use standard of IAA (0-50 µg/ml) for calculation the concentration in your Sample
Hi,
Do you mean auxin (IAA)? Here is a rapid method that I'm attaching. Hope it help you.
Cheers.
If you need a qualitative assay only, you can mix, on a filter paper, a drop of your sample with a drop of one of the following reagents that gives characteristic color in case of positive IAA production
1) FeCl3/perchloric acid reagent: ( 1 ml 0.5M FeCl3+ 49 ml 35% PCA +50 ml 96% ethanol), gives pink spot with IAA
2) DMCA reagent: (1 g dimethylamino cinnamaldehyde + 50 ml ethanol), gives purple spot with IAA
Well as rightly pointed out it is IAA and not AAI. Now, if have a pure compound of IAA which can used as primary standard then simply do HPLC with C-18 column of still better GC with PORAPAK -Q as the column with TCD or still better ECD as the detector by using the pure IAA as primary standard..
Hi
You can use spectrophometric method as following:
prepare Salkowski reagent (FeCl3 10mM /perchloric acid 35%)
- Centrifugation of bacterial culture suspension at 10000g for 10 min at 4°C
- Take 1 ml of free bacteria supernatant
- add 2 drop of orthophosphoric acid
- add 2 ml of Salkowski reagent
- Incubation at room T° for 20-25 min
in presence of IAA, we see change the color to pink
Take OD at 535 nm against a blank prepared in the same condition with bacterian medium
you can use standard of IAA (0-50 µg/ml) for calculation the concentration in your Sample
Hi
You can also performed a quality test as follows as initial step whether your strains have capacity to produce IAA
A loopful of test culture will be inoculated to the tryptone broth and incubated at 37 ̊C for 24 hours.3-4 drops of Xylene was added after incubation and it was shaked it vigorously and allowed the two layers to separate. 1ml of Ehrlich’s reagent was added slowly to form the layer on the surface of xylene. The formation of pink colour was observed at the lower surface of xylene layer (Rakesh patel.,et al 2006).
or
Bacterial isolates will be inoculated for determination of IAA like substances in 5ml of peptone water supplemented with Nacl media (Vendan et al. 2008). The indole strip will be put with overnight incubation at 37C. Next day if the stip is turn from yellow to pink colour than the test is positive.
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