basic doubts in mapping

[rathankar]

I have a doubt in mapping studies
I had taken a set of proteins and obtained the mRNA from NCBI and then converted them in to reads using a perl program. If i want to map them back to the reference sequence, I used BWA and I want to visualize the mapped file [sam format] using tablet viewer.

1. since I did a fasta to fastq conversion, and if the mRNA coding the hypothetical protein was expressed in the negative strand, will it be meaningful if i wish to map this mRNA [cDNA] to the reference sequence. Or in other words, should i also take the complement of this sequence and then convert it in to fastq format. pls let me know

2. When, I did a mapping of these sequences against the reference sequence, for each mRNA sequence, I got it mapped to 16 different locations in the reference sequence and in fact none of them were overlapping. This is what i understood by looking at the beginning and ending coordinates of each alignment in the SAM file.

3. I wished to view this mapping in a tablet viewer, but it shows an error everytime i load [java.source.lang not found]. Though other java programs runs effectively, I am unable to view the output.

4. If I want to find the promoters of one such protein, i thought of taking 200bp upstream to the beginning of the gene site and then get the corresponding coordinates. But is the prediction approach correct.

pls help

with regards
N. Rathankar

[Dario1984]

The strand is not relevant. You should not reverse complement it.

You did not provide enough detail about your RNA mapping. How long are your simulated reads ? If they are too short, they can map to multiple places in the genome. Also, if you didn't set the BWA options carefully, it can return mappings with different numbers of mismatches.

It is not clear from your description when the error occurs. Is it when you load the program, or does the program start fine, and give you an error only after you try and import your data ?

Determining promoters by your method is reasonable.