transcriptome sequencing help - (May/27/2011 )

Hi guys,i want to do transcriptome sequencing through next generation sequencing approach.But i am facing problems with it.Actualy i am confused from where to start.Plez suggest me how i should do this job, like from where i should start, which tools i should use.

You can start out by reading literature on deep sequencing of transcriptome. Many papers have been published recent years. Also there are different strategies of preparing samples for sequencing depending on the purpose of your study such as whether you focus on non-coding or small transcriptomes. Typically, RNA is isolated, cleaved into small spices, converted into cDNA followed by 2nd strand cDNA synthesis. the dsDNA is then end repaired, ligated to adaptors, amplified, and sequenced. Bioinformatics mapping and analysis of the data are then involved.

pcrman on Sun May 29 00:07:31 2011 said:

Dear bro,you are dead right.But the fact is i want to do it using bioinformatics tools from beginning.Actualy it has been a while now using bioinformatics tools.I have to only focus on bioinformatics tools.One more thing my purpose is to study small transcriptomes,and i want to do only using bioinformatics tools. For e.g,as for as i have understood,i have to look for the gene which would probably undergo alternate splicing.Then i have to pick cDNA of its different variants.from here,it becomes rather confusing for me to go ahead.Plez suggest how should i proceed by using only bioinfomatics tools