Systematic analysis of mRNA 5' coding sequence incompleteness in <it>Danio rerio</it>: an automated EST-based approach

• Main Authors: Zannotti Maria, Carinci Paolo, Facchin Federica, Vitale Lorenza, Canaider Silvia, Lenzi Luca, Casadei Raffaella, Frabetti Flavia, Strippoli Pierluigi
• Format: Article
• Language: English
• Published: BMC 2007-11-01
• Series: Biology Direct
• Online Access: http://www.biology-direct.com/content/2/1/34

<p>Abstract</p> <p>Background</p> <p>All standard methods for cDNA cloning are affected by a potential inability to effectively clone the 5' region of mRNA. The aim of this work was to estimate mRNA open reading frame (ORF) 5' region sequence completeness in the model organism <it>Danio rerio </it>(zebrafish).</p> <p>Results</p> <p>We implemented a novel automated approach (<it>5'_ORF_Extender</it>) that systematically compares available expressed sequence tags (ESTs) with all the zebrafish experimentally determined mRNA sequences, identifies additional sequence stretches at 5' region and scans for the presence of all conditions needed to define a new, extended putative ORF. Our software was able to identify 285 (3.3%) mRNAs with putatively incomplete ORFs at 5' region and, in three example cases selected (<it>selt1a</it>, <it>unc119.2</it>, <it>nppa</it>), the extended coding region at 5' end was cloned by reverse transcription-polymerase chain reaction (RT-PCR).</p> <p>Conclusion</p> <p>The implemented method, which could also be useful for the analysis of other genomes, allowed us to describe the relevance of the "5' end mRNA artifact" problem for genomic annotation and functional genomic experiment design in zebrafish.</p> <p>Open peer review</p> <p>This article was reviewed by Alexey V. Kochetov (nominated by Mikhail Gelfand), Shamil Sunyaev, and Gáspár Jékely. For the full reviews, please go to the Reviewers' Comments section.</p>