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I am in the process of assembling and annotating the genome of a non-model organism, using almost exclusively short read (paired-end Illumina) data. Throughput is one obvious benefit of these data (high coverage for reasonable cost, error rates notwithstanding), but another benefit of RNA-seq data in particular is that they can be used in multiple ways. For example, I am mapping the RNA-seq reads to the genome to estimate transcript abundances and do some differential expression analysis, but I have also assembled the RNA-seq reads de novo to use as evidence for genome annotation.

There are, however, limitations to using RNA-seq-derived transcripts for annotation. Full-length cDNAs would facilitate much more reliable annotation. However, I have no intuition for the relative cost, time commitment, or complexity of extracting and sequencing full-length cDNAs vs next-gen sequencing. Can anyone here comment on this?

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What kind of organism? Splicing makes using RNA more difficult. Is there a good existing reference genome for you organism (if lacking in annotation)? Do you have a specific goal, such as biofuels, drug production fighting pathogenic or parasitic organism, or do you have X amount of money and want to do the best basic research? The type and depth of sequencing that is most cost efficient will vary with your goals. –  Stuart R. Jefferys Nov 29 '12 at 20:29
"What kind of organism?" It's eukaryotic, a social insect to be precise. "Is there a good existing reference genome for you organism?" As I said, this is a non-model species, so I am the one assembling the genome. I don't know what your criteria are for a "good" genome: no physical mapping, gap filling, etc has been done yet, but the assembled sequence is close to the expected genome size and preliminary annotations have identified about the number of genes we would expect (perhaps a little bit less). –  Daniel Standage Nov 29 '12 at 21:25
"Do you have a specific goal?" The PI of this genome project is primarily interested in the intersection of evolution, genomics, and behavior, and uses several species of social insects as model organisms for these subjects. –  Daniel Standage Nov 29 '12 at 21:27

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