Hello,
Would any one please tell on which samples the digestion is occuring? the products are 107 and 82bp..
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#2
pito
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Posted 25 July 2010 - 06:08 AM
Its the same gel, but just 2 pictures?
And you see a band near 100bp in every lane... but is this digested?
Where is your negative control? (the sample without digestion?)
And you see a band near 100bp in every lane... but is this digested?
Where is your negative control? (the sample without digestion?)
If you don't know it, then ask it! Better to ask and look foolish to some then not ask and stay stupid.
#3
Fhannan
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Posted 25 July 2010 - 07:43 AM
No these are different samples..
I dont load a -ve control as the presence of only the 107 bp without the 82bp should work as -ve control...
i presume the last four wells of pic 1 are not showing any digestion and should be considered as the -ve control..
my suspects are the shadow bands, do they represent a +ve reaction where digestion is taking place???
I dont load a -ve control as the presence of only the 107 bp without the 82bp should work as -ve control...
i presume the last four wells of pic 1 are not showing any digestion and should be considered as the -ve control..
my suspects are the shadow bands, do they represent a +ve reaction where digestion is taking place???
#4
pito
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Posted 25 July 2010 - 07:55 AM
Fhannan, on 25 July 2010 - 07:43 AM, said:
No these are different samples..
I dont load a -ve control as the presence of only the 107 bp without the 82bp should work as -ve control...
i presume the last four wells of pic 1 are not showing any digestion and should be considered as the -ve control..
my suspects are the shadow bands, do they represent a +ve reaction where digestion is taking place???
I dont load a -ve control as the presence of only the 107 bp without the 82bp should work as -ve control...
i presume the last four wells of pic 1 are not showing any digestion and should be considered as the -ve control..
my suspects are the shadow bands, do they represent a +ve reaction where digestion is taking place???
If you did load a negative control... you would not have this problem.
And the be honest: I see no difference in pic 1 between the last 4 and first 4 lanes.. But this can be due the low quality of the pic I am seeing here when I zoom in.
and what is a positive controle?? You allready have a sample that has been digested before???
In the +ve lane, I hardly see anything at the +-100bp ?
If you don't know it, then ask it! Better to ask and look foolish to some then not ask and stay stupid.
#5
perneseblue
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Unlimited ligation works!
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Posted 25 July 2010 - 08:40 AM
Sorry separation on this gel isn't good enough. What kind of gel are you using?
May your PCR products be long, your protocols short and your boss on holiday
#6
Fhannan
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Posted 26 July 2010 - 12:16 AM
12% PAGE, the lower bands are primer dimmer...my question is should the digested bands appear as intense as the undigested band on the gel? or i can consider the faint band as the lower digested product?
