I got troubles that I PCRed certain genes from the Arabidopsis phage cDNA library (from Arabidopsis Biological Resource Center ) these days, but all of the results of PCRs only displayed a few nonspecific amplifications. All of my target genes are 2000bp+, however the nonspecific bands gathered in 200-500bp.
Now, should I make sure of these things at first?
1.The quality of cDNA, because it was kept in -80 for 2 years+ and not used before. Is it necessary to use beta-actin to test the cDNA?
2.The concentration of cDNA template is unknown, so I'm no sure if the content of template I added(2ul cDNA for 50ul) is below the requirement. Could the presence of nonspecific bands revealed the template content is enough in fact?
Need for your suggestion, thanks so much
PCR from cDNA library
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