DNA from peripheral blood (isolated by classical or kit methods) represent both WBC and cancer cells. If gene X was a tumor suppressor that can be inactivated in cancer, it is possible to assume that unmethylated Band comes from WBC and the methylated band comes from cancer cells. But this is very general rule.

In my experience there are exceptions to this rule. For example, we analyzed CFTR and p16 genes and they were partially methylated in a number of normal control samples.

It is logical to say that but I never come across such suggestion in any articles. would you provide me any references ?

ok.. then, based on my sequencing result in the attached PDF does it mean that the sample is partially methylated ?

yes right Yazan Haddad 

AMIRA : DNA from peripheral blood sample (isolated by classical or kit methods available from market) normally represent both WBC and cancer cells(basic). If gene X was involved in  a tumor suppressor and inactivated in cancer. Then  it is possible to assumed  that unmethylated Band comes from WBC and the methylated band comes from cancer cells.But  few  exceptions cases  CFTR and p16 genes and they were partially methylated in a number of normal control samples. In this cases it partially

methylated. My suggestion is same time you little focused on the gel equipments negative and positive charge,and also make another experiment in your friends

gel system or other equipment. I am assuring you you will finally observed clearly

what are the mistake going on this experiment please actively and be cool mood

handle the experiment once again and confirmed with both experiment.

Definitely your getting correct ans ware for the same it happens many times in wet lab.

Thanking you and my best wishes 

very nice explanation dear Mayadhar Barik · 

Thanking you Sir

Thank you everyone for your kind help.

I will further discuss all the information with my colleagues and supervisors. 

I welcome more information regarding this issue.

Thanks again.

Well come

Your experiment is lacking important controls--or they are not shown on the gel.  These controls would be the inclusion of 100% methylated DNA, usually prepared by treating DNA with CpG methylase and 100% unmethylated DNA which can be prepared in various ways, including whole genome amplification or the use of certain mutant cells which lack DNA methylase.  With these two different control DNAs, you would bisulfite treat them and run your methyl specific PCR alongside your samples.  They should show the proper amplification pattern, demonstrating that your M and UM specific primers have the expected specificity with the PCR conditions that you are using.  Depending on the design of the MSP primers, the exact PCR conditions can be quite critical.  In many papers where partial methylation is measured, the authors will then use appropriate mixtures of the control methylated and unmethylated DNAs to simulate various possible levels of partial methylation.  For instance 5%M 95%U, 20%M80%U, 50%M50%U, 80%M20%U, 95%M5%U or other intermediate combinations.   They will then compare the patterns that they see in their sample of interest with their artificial standards and make some conclusion about the level of partial methylation.  Although this type of approach will give you some information, it is probably not as informative as bisulfite sequencing, which would provide more specific information about what is happening at each methylatable site.