HTSeq-count (Unexpected result of .sam and .gtf)

Question: HTSeq-count (Unexpected result of .sam and .gtf)

3.8 years ago by

Singapore

jchen015 • 80 wrote:

Dear all

I am trying to map the readings between a .sam file and a .gtf file by leveraging on the functionality of HTSeq-Count. However, there are some unexpected result which i presume the problems lies with the format of the files.

.sam file format : https://www.dropbox.com/s/r226fsejimmey3j/sam_data.png?dl=0

.gtf file format : https://www.dropbox.com/s/wfjrgraw8o6r33i/Human_gtf.png?dl=0

downloaded at (http://asia.ensembl.org/info/data/ftp/index.html?redirect=no)

results of the map result : https://www.dropbox.com/s/ls4atwyrwsri4t8/result_for_cut_L7_1_5_sam.png?dl=0

The result shows that the readings are:

no_feature:1511

.....

not_aligned: 22998

I am really not expecting this result, may I know where am i wrong at?

Regards,

Julius

gtf sam htseq-count • 1.4k views

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modified 3.8 years ago • written 3.8 years ago by jchen015 • 80

3.8 years ago by

Hotz, Hans-Rudolf • 1.8k

Switzerland

Hotz, Hans-Rudolf • 1.8k wrote:

I am not an HTSeq-count expert at all, but I have a guess:

As far as I can see, your sam file picture only shows reads which do not map (i.e. sam flag is 4). It might be helpfull to show a line with a read which is mapping to a chromosme, in order to check the format of chromosome names (e.g. "chr1" or "1").

Just in case it is "chr1", you most likely run into troubles, since the chromosome are labeled "1","2", etc in your gff file

Hans-Rudolf

ADD COMMENT • link written 3.8 years ago by Hotz, Hans-Rudolf • 1.8k

Hi hans-rudolf

Thanks for the reply. What if I require a mapping of my sam files to produce a visualization of the genome, genes and short reads, what should be the appropriate way of achieving it?

If it is impossible, then what are some recommendations you would think I should adopt to resolve this issue?

Regards,

Julius

ADD REPLY • link written 3.8 years ago by jchen015 • 80

Hi Julius

I am struggling to understand your latest comment.....are you still talking about your HTSeq-count problem? If you have a differnt question (about visualization?), please start a new thread.

Hans-Rudolf

ADD REPLY • link modified 3.8 years ago • written 3.8 years ago by Hotz, Hans-Rudolf • 1.8k

Hi Hans-Rudolf

My apologies.

Let me rephrase my comment. What I am trying to say here was......

I am unable to modify my .sam file, that is the format that I received from my source. My concern now is, if I want to map the readings for this .sam file, how can I do that? Am I able to do it with a gtf file ?

Referencing to your comment: "It might be helpfull to show a line with a read which is mapping to a chromosme, in order to check the format of chromosome names (e.g. "chr1" or "1")."

How can I do that?

Regards,

Julius

ADD REPLY • link written 3.8 years ago by jchen015 • 80

sorry, but it is just getting more confusing.....and I start wondering where is the connection to Galaxy?

Anyway, you have a sam file. sam files are tab delimted text files, hence you can open your file in any text editor, and of course you can use the 'Text Manipulation' and 'Filter and Sort' tools in Galxy to look at it. I recomend reading up on sam file specification to understand what each cloumn means.

Hans-Rudolf

ADD REPLY • link written 3.8 years ago by Hotz, Hans-Rudolf • 1.8k

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