I successfully amplified fungal ITS from soil samples, however after running the purified PCR products in an agarose gel they are barely visible and don't look like defined bands but rather clouds. The purification was done with the Monarch PCR and DNA Cleanup Kit. Why could this be happening?
PCR purification columns do cause a large loss of product. You can get a better yield by washing the dna off the column with hot 70c water or elution buffer but depending on your next process you may want to exo-sap the product to get rid of primers or just ethanol precipitate the pcr product for later use. Many later stages can take place in pcr buffers ( like restriction digests) so you may want to check what level of purification your samples need
Hello,
If a gel pic can be uploaded, the problem can be visualized easily
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