The seeds I got from NASC have no kanamycin resistance despite having the NPTII gene/T-DNA insertion. I tried growing seedlings on 5'-aza-2'-deoxycytidine to reset any methylation related silencing. However, with no succes.
Can anyone suggest a way to regain kanamycin resistance?
thanks!
I also faced similar problem and I hope we can get any suggestions for it
Unfortunately a lot of NASC seeds have lost their kanamycin resistance, I have not heard of anyone getting it back. I would suggest a quick DNA extraction and genotype these lines by PCR.
http://www.plantmethods.com/content/1/1/4 - we have found that the sucrose method works without grinding or shaking, so very quick and efficient.
We have observed that some progeny of gene-silenced GFP lines become expression after they are backcrossed to the wild-type (try reciprocal crossing). Not sure why, but we know that the 'methylation' marks suppose to be wiped out during 'zygote' stage, and later reset again.....
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