The way you posted question here, it can be interpreted that you might have done some mistake. Also, the sequencing was not good. So there were many problems. I suggest you to discuss the problem with someone in your lab who is more experienced in this kind of analysis.

Nevertheless, you can also send them your amplicon primers, if M13 primers were not working good.

But you DNA is from pure culture or from soil? Sovetgul Asekova

The peak spacings are ok but the peak intensity is in the range90-108 while the background is 7-8 so the signal/noise ratio is very poor. The reaction runs the full length of the amplimer but it is clear that there are 2 sequences or 2 sequencing primers present. I will assume that the primers were purified away from the pcr product with sephadex columns from the .abi file information.

If we assume that your pcr product was a single pure band then some ways of generating a result of this kind are

1 Not enough template so low intensity peaks.....does not account for the messiness of the electropherogram

2 not enough sequencing primer....again electropherogram is too dirty.

3 2 primers (sequencing) present possibly the non m13 and the m13 primer both sequencing from a different position at the same time in the same direction or even one primer from each end sequencing simultaneously by mistake

4 both primers (sequencing and pcr) have the same m13 sequence so the product is being sequenced from both ends at the same time

What was your template purification method and what were the pcr and sequencing primer sequences please?

Abhijeet Singh, Paul Rutland, Giorgia Pertile my gratitude for early responses

Abhijeet Singh I have done this experiment a couple years ago, I might do some mistake apparently..and here to mention yes, I sent my amplicon primers of 10pmole as well... When I asked one of my colleague he suggested me to use different Taq enzyme . I used from Solgent's e-Taq and rxn reagents. Giorgia Pertile this samples are from Plant tissue extracted using by standard CTAB method adding RNAsy treatment, my DNA concentration is okay I think since I extracted many DNAs and amplification by PCR predominantly were successful. The below attached image is PCR amplification checkup using my gene-specific designed primer on QIAXCEL electropherogram. The bands have appeared clean which is why I directly sampled and sent to the sequencing company with the preference to do "Purification for unpurified PCR product". Paul Rutland Sir, from your above-mentioned #3 and #4 comment points make me think and this time I shouldn't have ordered m13 forward and reverse primer involving to my sequencing samples of 200 bps. I designed two gene-specific primers within one targeted gene of disease resistance "R gene' (first is 200 bp which I sent for sequencing and another next in line is second primer of ~600 bp in product sizes) The experiment I am doing is running out of time, I would like to hear your opinions again on my next plan as I would like to send another PCR product of ~700 bp (below I attached picture, second screenshot). Those "E4", "E5" and "E7" are acceptable for sequencing or should i change my Taq enzyme and run on standard gel then cut the gel and send them? Thank you for your valuable advice

Ok but the DNA is from one type of plant or more...? Because your results is same when happear more polimorphism inside your sequence.

Dear Giorgia Pertile thank you for your comment

it is only Plant DNA from 3 different cultivars

the bands look clean so I would cut them out and purify and send the dna and the original pcr primers as sequencing primers both forwards and reverse to ensure good coverage at the ends of the sequence Sovetgul Asekova .

Paul Rutland I sincerely appreciate your guidance, thank you