Protocol Online logo
Top : Forum Archives: : Molecular Biology

Can anyone suggest any paper,journal or book to read about analysing sequenced r - (Apr/22/2008 )

blush.gif I know this must be quite stupid...but better admit it than sorry, right? I have difficulties understanding sequenced result. wacko.gif I also find it hard for me to do further analysis with the sequenced result, due to less understanding of it. It would be great if any kind soul here would like to guide, teach, suggest anything that can be used to brush up my stupidity in this. Thanks in advance.

Cheerioet.

-cheerioet83-

Do you have a software to read your sequencing?

Few key thing you should know: A good sequence will have well separated ACTG peaks. At initial peaks, it is normal to have not so nice peaks and also at the last few peaks.

Just google on sequencing, or download some software. I cant recall the name of the software, when I found out, I will let u know.

-timjim-

Sure..thanks for your help. I will wait for your good news, then. I had also asked Val about it...It's just that I didn't understand it well. tongue.gif
If I had Blast it at NCBI and what should I look for? The nearest percentage to perfect?

Thanks in advance. rolleyes.gif

-cheerioet83-

chromas software is eay to use but offers not too many features for analysing electropherograms.
Other than that I've used parts of the Vector NTI package from invitrogen (free for academic users).

for more info on blasting, i suggest you read the tutorials on NCBI: http://www.ncbi.nlm.nih.gov/Education/BLASTinfo/tut1.html and http://www.ncbi.nlm.nih.gov/Education/BLASTinfo/tut2.html

-vairus-

Oh...thanks for the infos...I will look into it..and if I have quiries...I will sure ask again at here.. biggrin.gif

-cheerioet83-

A common problem with most people who start working with sequence is to look at the returned sequence, but to ignore the electropherogram. Until you are familiar with sequencing, I would ignore the returned sequence, delay blasting, and spend time looking carefully at your electropherogram. Chromas or 4Peaks(Mac) or Vector NTI will allow you to easily look at the peaks. You will see for yourself where the difficulties are, how the program is having trouble (or not) reading sequence, and have the ability to see if the problem is your sample or the basecaller. You can also optimize your sample preparation to give good sequence, a step often omitted. You will know that the first 20 bp are usually garbage, that dye blobs usually happen at 80 and 120 bp, and that repeated bases after 850 bp are likely to have the number of bases wrong. None of these things are possible to see if you only look at the called bases.

-phage434-

Thanks Phage....I had indeed downloaded Chromas and viewed the sequence with it.

-cheerioet83-