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| kitty |
 Posted: Fri Apr 25, 2008 7:26 pm Post subject: MICROBIOLOGY |
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Forum Freshman
Joined: 28 Oct 2007
Posts: 11
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HI!
I need help on this...I don't understand how can to interpret the results of osmotic shock in a whatever bacteria, using for example, B-lactamase as indicator, in the three steps of osmotic shock: EDTA, hypertonic solution and hipotonic solution, if I don't know where the enzime is located ( cytoplasm or periplasm)
I know this experiment is used for this, but I don't know how to interpret the results: if is positive o negative ( the location of enzime) in each of the step, and if is citoplasmic or periplasmic.
please some body could help me!
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| i_feel_tiredsleepy |
| Posted: Sat Apr 26, 2008 8:57 am Post subject: |
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Forum Senior
Joined: 21 Mar 2008
Posts: 376
Location: Montreal
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Ok I'm not exactly sure which experiment you did but.
EDTA disrupts the integrity of the outer-membrane, so I guess if your indicator is still present after treatment with EDTA then it is cytoplasmic.
Is this the one where you treat with lysozyme to see if it will lyse? Then you get:
Just lysozyme hypertonic: No lysis
Lysozyme + EDTA: Lysis
Which shows that the cell wall gives structural integrity to the cell.
If it's something else I've never heard of the experiment lol. |
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| kitty |
| Posted: Sat Apr 26, 2008 9:58 am Post subject: =( |
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Forum Freshman
Joined: 28 Oct 2007
Posts: 11
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The experiment consist to know where is the enzyme(B-lactamase) located( in the periplasm or cytoplasma) so, the bacteria is treated with osmotic shock, 3 steps: EDTA, hipertonic solution and the hipotonic solution. Before this, is necesary to centrifugate and finally I have a pellet and supernatant ( of each step) in wish add a indicator ( Nitrocefina).
And my doubt is...what results I expect to see in each step?
in where place I expect to see color of the indicator if is located in periplasm(or cytoplasm) in the 3 steps: in the pellet or supernatant?
example: the enzyme is located in cytoplam
steps 1, EDTA: supernatant (color or not color?)
pellet (color or not color?)
step2, hypertonic: supernatant(?)
pellet(?)
step3, hipotonic: supernatant (?)
pellet(?) |
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| i_feel_tiredsleepy |
| Posted: Sat Apr 26, 2008 10:03 am Post subject: |
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Forum Senior
Joined: 21 Mar 2008
Posts: 376
Location: Montreal
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| I posted my response in your other thread lol. |
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