The same set of PCR conditions including the cDNA tube once gave nice clear bands, but two days later there were absolutely no bands at all (i have repeated this twice). This pattern has occurred some three times till now.
Am absolutely clueless as to what is happening.
I hope you guys help me out
Hi Ramya,
There are many parameters that you might need to consider for no amplification:
1. You might want to consider using a fresh stock of Primers and repeat the PCR. There are chances where Primers tend to degrade. If the primers
2. Use a fresh cDNA sample and repeat it again.
3. Autoclave and use your tips and tubes again.
Hope this is of some use to you!
All the best!
Thanks ...ll try option 1 as i regularly autoclave tips and tubes i use
Are you using DNase free water to dissolve your primers and cDNA stocks and other PCR components.
Some times cDNAs tend to degrade by repeated freeze thaw.. Try fresh one.. and as mentioned above.. all reagents new.. Primers/dNTPs/ and especially Water..!! May be there are some nucleases which tend to degrade your cDNA
I got it solved people.
I have started to increase the enzyme concentration
i dont know whether it is the more enzyme or the glycerol that has made the reaction worked :)
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