I have extracted bacterial genomic DNA from CTAB method and done an agarose gel electrophoresis to check the quality. Then I got clear DNA bands (image). PCR was done using 16s rDNA primers. (1.5Kb) When the PCR Product was gel electrophoresed, I could not get the 1.5 kb band from them except for 2 samples. (These two also were very thin light bands). I purified those two and again ran a gel. Only one gave a band. Can someone help me with this?
It would be interesting to see some samples from the gel you show but run with a dna ladder to get some idea of size. The gel image looks like a lot of either rna or degraded dna except for samples 4 and 11 on the top row. Is it possible that your samples degraded before genomic dna purification. Which 2 samples worked and gave a pcr amplimer?
Dear Mr. Rutland,
Thank you. The attached image was the genomic DNA just after extraction.I’ll upload the PCR product image also
Hi Kalani!
I am not an expert of the CTAB method you commented for DNA extraction. However, I would like to emphasise the use of proteinase for a proper cell lysis and as Paul commented previously, given the low intensity of the band the DNA may have undergo degradation (samples 4,5,11..) or simply low yield of the process
I hope this helps!
Good day!
In my opinion, the absence of bands on the gel the amplicons were viewed on after PCR may indicate degradation of buffer or primer used
It may also indicate contamination of stock sample
- Badges
- Science topic
- Similar topics
- Chemistry, Organic
- Organic