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sequencing result doesn't match my gene.... - (May/09/2012 )

Hi all,

the project that i am currently doing is about the genotyping of SNP (human) by using PCR-RFLP. i am facing some problems with the sequencing result (using PCR products of 200 bp):
1) My sequencing result doesn't show high % identity with the gene that i am studying but show high identity with other gene after i Blastn the result.
2) in the result, the sequence should contain the primers and SNP right? but i couldn't locate any of them in the result that i obtained.. how could this happened?
3) the sequencing chromatogram doesn't show any multiple/ overlapping peaks.. the peaks are regular and nicely separated.

As the purpose of my study is to genotype the SNP and confirm it by performing sequencing, the sequencing result that i obtained doesn't appear to be as what i expected...kinda frustrated..

any idea? anyone can plz help me?

tq very much!


Hello Micki :)

make sure of your primers ...


Do a primer blast (NCBI) and check the possibility of amplifying your target and the suspect....


my primers supposed to amplify the sequence that i want. but in my sequencing result, the primer sequence even didn't appear in it.. is it possible to get such result?

any idea about this?


the length of my sequencing result is even longer than the pcr product length... very confuse...


You probably have someone else's sequencing results. Or your samples were mixed up prior to sequencing.