Making normalized cDNA libraries

[m.nyine]

Dear Members,

I am a fresh graduate student who is interested in bioinformatics. My work requires me to generated a reference transcriptome from mRNA. Help me understand some basic concepts of how to generate a normalized cDNA library.

What is paired end sequencing as required to make scafolds from contigs?

Thank you.

NM

[grassgirl]

NM,

A normalized cDNA library makes it so that the highly-expressed genes, such as housekeeping genes, don't overwhelm your output and you can find other genes that aren't as highly expressed more easily. The company Evrogen has a kit called Trimmer for normalizing. Look the company up and contact them about what platform you are using, etc. and they should be able to point you to the right kit and protocol.

As for paired-ends and cDNA libraries, it depends on the platform again. People do this for the Illumina, which has short reads, but the 454 Titanium has longer reads and paired-end isn't necessary.