After DNA isolation and PCR of my endophytic fungal isolate, neither the gDNA nor the PCR product showed up on the gel but when tested on the nanodrop both displayed high concentrations of DNA. I initially thought this was due to the nanodrop picking up fragments of degraded DNA but when I sent this PCR product for sequencing the isolate was successfully sequenced and identified as an Aspergillus japonicus clone. Does anyone have any likely explanations as to why this isolate would not show up on the electrophoresis gels ?
It depends a lot on the size of the gel comb and the dna dye that you use in your gel and the distance that you ran the pcr product in the gel
The amount of pcr template used for sequencin is
100–200 bp 1–3 ng
200–500 bp 3–10 ng
500–1000 bp 5–20 ng
but the minimum amount of dna that you can see on an ETBR stained gel is 10ng and if the wells are large then this could easily be 20-30 ng so it is possible to have a band containing 20ng of dna which will not be visible especially if it has been run a long way and diffused in the gel but still enough to sequence
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