Not able to amplify the gene segment which showed a positive result earlier. Tried three different protocol along with master mix still no result?

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Craig S Wilding

Are you working with cDNA or genomic DNA? If you got a positive result earlier and are working with cDNA, but now fail to amplify the fragment then it may be that your cDNA is degraded. Defrost a new aliquot and try again.

Sudeshna Thakur

I am working with cDNA and yes I have now freshly synthesized the cDNA to work with.

Thank you for the comment.