Natarajan et al. BMC Genomics 2010, 11:606 http://www.biomedcentral.com/1471-2164/11/606 submitted for functional classification using GO annota- tion tool (GOSlim) at TAIR. Functional classification was done under three Gene Ontology categories viz., cellular component, molecular function and biological process. These three broad categories were further clas- sified with different GO Slim terms. Semi-quantitative RT-PCR Seventeen ESTs for oil biosynthesis genes and one EST for actin gene were selected from the current study and sequenced from 3’ ends. Using these 3’ end sequences, primers were synthesized to amplify the 3’ UTR exclud- ing the poly-(A) tail for all the genes except actin. For actin, the primers were synthesised to amplify a 590 bp fragment which included 392 bp from the 3’ end of the coding region and 198 bp from the 3’ UTR. The primer sequences and the size of the amplified fragments are given in Table 3. For RT-PCR, total RNA from roots, mature leaves, flowers and developing seeds were trea- ted with DNase and purified using RNeasy Mini Kit fol- lowing the manufacturer’s protocol (Qiagen, Hilden, Germany). About 3.0 μg of purified total RNA from each sample was used for first strand cDNA synthesis using 50 pmol oliog-dT(18) primer and 100 units of Pri- meScriptTM reverse transcriptase (Takara Bio Inc, Shiga, Japan). Equal quantity of first strand cDNA (from 25 ng total RNA) was used for PCR. Semi-quantitative analysis of the PCR amplified fragments was done by agarose gel electrophoresis and ethidium bromide staining. Accession numbers J. curcas unigenes were submitted to the dbEST database of Genbank at NCBI with the accession numbers GW874611 to GW881590 and HO004465 to HO004493. Acknowledgements Financial support provided for this project by the Department of Biotechnology, Government of India (Sanction order BT/PR/8647/PBD/26/38/ 2007) and the 16-capillary automated DNA sequencing machine (Genetic Analyzer 3130xl, Applied Biosystems) provided by the SRM University are gratefully acknowledged. Authors’ contributions The study was conceived and directed by PM. RNA isolation, construction of normalized cDNA Library, plasmid DNA preparation, automated DNA sequencing, assembly, annotation, validation of ESTs and other bioinformatics analysis were directed by PM and carried out by PN. Transformation and patching clones were done by DK and PN. Plasmid DNA Isolation and annotation was assisted by GG, JP, NS, PAS, and KKS. PM and PN wrote the paper. All authors read and approved the final manuscript. Received: 19 June 2010 Accepted: 27 October 2010 Published: 27 October 2010 References 1. Mabberley DJ: The Plant Book, A portable dictionary of vascular plants. 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