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Help on DNA quantification and RAPD...

Naresh C. Pancholi N.C.Pancholi at reading.ac.uk
Wed Jul 27 08:36:47 EST 1994


Hello Netters,
I would appreciate any help on the following questions:

1. I am using CTAB DNA extraction method for banana leaves.  I tried to 
quantify DNA through UV spectrophotometer but this reading is not reliable as 
CTAB also absorbs fluorescence.  Then I tried fluorimeter but I am still
not happy.  It seems that my samples have very low DNA because my starting leaf
material is only 50-70 mg.  Can someone tell me a reliable, quick and cheap
method for DNA quantification?

2. On the basis of this, I am going to use RAPD and I have a big sample size
(300 plants) and less time (2 months only).  I use primer kits from Operon.
I noticed a strange phenomenon in my RAPD reaction.  I tried 10, 25 and 50 ng
of template DNA (based on fluorimeter readings) per 25 ul reaction.  There was
an increase in the total number of bands as the concentration of DNA increased.
What I understand that this should not happen, though I am a novice for this
technique and therefore, would like to receive your advice on this matter.

3. Is any one there who is comparing a big sample size like mine, through
RAPD?  If so, would you please write general or any specific advice for it?

I would appreciate any feedback on this matter from you.  I have also posted
this message to Methods group so sorry if you have come across again. Thanks
a lot in advance.

+-----------------------------------------------+
|  Naresh Pancholi       abrpanch at reading.ac.uk |
|  Agricultural Botany   Phone: +44 734 318092  |
|  Reading University    Fax:   +44 734 316577  |
|  Reading RG6 2AS                              |
|  UK                                           |
+-----------------------------------------------+    





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