Hello everybody,
we have a trouble about staining of ISSR PCR products. we have PCR products but staining is problematic. We tried several things but we couldn't succeed. Is there anyone who has a protocol to solve this problem please? Thank you everybody.
Have you tried run the ISSR product on polyacrylamide gels, and stain the gel with Silver-staining method. Silver-staining method should be more sensitive. I attached this paper for you. See below.
Why you are doing staining by using EtBr in agarose gel. First confirm your PCR product whether it is amplifying or not?
Good luck...
I am having a hard time identifying any amplification on 1.5% agarose with gel Red. Same problem.
@ Madeline,
If you stain your gel with GelRedTM, it is better you stain it post-electrophoresis. The dye can modify the migration of your DNA fragments on gel. See this discussion on RG:
https://www.researchgate.net/post/Does_anybody_know_why_GelRed_modifies_the_migration_of_DNA_fragments_in_agarose_gel_electrophoresis
For ISSR PCR product profing you can use 1.4% of Agarose gel and also use ETBR as a staining dye but take from the stock concentration of 1mg /1ml
eg. Preparation of 100ml , 1.4% Agarose gels.
Agarose-1.4mg
TBE buffer(20X)-5ml
DDH2O-95ml
ETBR-5ul
Add ETBR finally when we pour the agarose solution to the gel tray, Since ETBR is carcinogenic and avoid breathing vapours coming from the beaker while adding ETBR.
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