De novo assembly and transcriptome analysis of five major tissues of Jatropha curcas L. using GS FLX titanium platform of 454 pyrosequencing

<p>Abstract</p> <p>Background</p> <p><it>Jatropha curcas </it>L. is an important non-edible oilseed crop with promising future in biodiesel production. However, factors like oil yield, oil composition, toxic compounds in oil cake, pests and diseases limit its commercial potential. Well established genetic engineering methods using cloned genes could be used to address these limitations. Earlier, 10,983 unigenes from Sanger sequencing of ESTs, and 3,484 unique assembled transcripts from 454 pyrosequencing of uncloned cDNAs were reported. In order to expedite the process of gene discovery, we have undertaken 454 pyrosequencing of normalized cDNAs prepared from roots, mature leaves, flowers, developing seeds, and embryos of <it>J. curcas</it>.</p> <p>Results</p> <p>From 383,918 raw reads, we obtained 381,957 quality-filtered and trimmed reads that are suitable for the assembly of transcript sequences. <it>De novo </it>contig assembly of these reads generated 17,457 assembled transcripts (contigs) and 54,002 singletons. Average length of the assembled transcripts was 916 bp. About 30% of the transcripts were longer than 1000 bases, and the size of the longest transcript was 7,173 bases. BLASTX analysis revealed that 2,589 of these transcripts are full-length. The assembled transcripts were validated by RT-PCR analysis of 28 transcripts. The results showed that the transcripts were correctly assembled and represent actively expressed genes. KEGG pathway mapping showed that 2,320 transcripts are related to major biochemical pathways including the oil biosynthesis pathway. Overall, the current study reports 14,327 new assembled transcripts which included 2589 full-length transcripts and 27 transcripts that are directly involved in oil biosynthesis.</p> <p>Conclusion</p> <p>The large number of transcripts reported in the current study together with existing ESTs and transcript sequences will serve as an invaluable genetic resource for crop improvement in jatropha. Sequence information of those genes that are involved in oil biosynthesis could be used for metabolic engineering of jatropha to increase oil content, and to modify oil composition.</p

The First Illumina-Based De Novo Transcriptome Sequencing and Analysis of Safflower Flowers

BACKGROUND: The safflower, Carthamus tinctorius L., is a worldwide oil crop, and its flowers, which have a high flavonoid content, are an important medicinal resource against cardiovascular disease in traditional medicine. Because the safflower has a large and complex genome, the development of its genomic resources has been delayed. Second-generation Illumina sequencing is now an efficient route for generating an enormous volume of sequences that can represent a large number of genes and their expression levels. METHODOLOGY/PRINCIPAL FINDINGS: To investigate the genes and pathways that might control flavonoids and other secondary metabolites in the safflower, we used Illumina sequencing to perform a de novo assembly of the safflower tubular flower tissue transcriptome. We obtained a total of 4.69 Gb in clean nucleotides comprising 52,119,104 clean sequencing reads, 195,320 contigs, and 120,778 unigenes. Based on similarity searches with known proteins, we annotated 70,342 of the unigenes (about 58% of the identified unigenes) with cut-off E-values of 10(-5). In total, 21,943 of the safflower unigenes were found to have COG classifications, and BLAST2GO assigned 26,332 of the unigenes to 1,754 GO term annotations. In addition, we assigned 30,203 of the unigenes to 121 KEGG pathways. When we focused on genes identified as contributing to flavonoid biosynthesis and the biosynthesis of unsaturated fatty acids, which are important pathways that control flower and seed quality, respectively, we found that these genes were fairly well conserved in the safflower genome compared to those of other plants. CONCLUSIONS/SIGNIFICANCE: Our study provides abundant genomic data for Carthamus tinctorius L. and offers comprehensive sequence resources for studying the safflower. We believe that these transcriptome datasets will serve as an important public information platform to accelerate studies of the safflower genome, and may help us define the mechanisms of flower tissue-specific and secondary metabolism in this non-model plant

Global Analysis of Gene Expression Profiles in Developing Physic Nut (Jatropha curcas L.) Seeds

Background: Physic nut (Jatropha curcas L.) is an oilseed plant species with high potential utility as a biofuel. Furthermore, following recent sequencing of its genome and the availability of expressed sequence tag (EST) libraries, it is a valuable model plant for studying carbon assimilation in endosperms of oilseed plants. There have been several transcriptomic analyses of developing physic nut seeds using ESTs, but they have provided limited information on the accumulation of stored resources in the seeds. Methodology/Principal Findings: We applied next-generation Illumina sequencing technology to analyze global gen

Production of very long chain polyunsaturated omega-3 and omega-6 fatty acids in plants

We report the production of two very long chain polyunsaturated fatty acids, arachidonic acid (AA) and eicosapentaenoic acid (EPA), in substantial quantities in a higher plant. This was achieved using genes encoding enzymes participating in the ω3/6 Δ8-desaturation biosynthetic pathways for the formation of C20 polyunsaturated fatty acids. Arabidopsis thaliana was transformed sequentially with genes encoding a Δ9-specific elongating activity from Isochrysis galbana, a Δ8-desaturase from Euglena gracilis and a Δ5-desaturase from Mortierella alpina. Instrumental in the successful reconstitution of these C20 polyunsaturated fatty acid biosynthetic pathways was the I. galbana C18-Δ9-elongating activity, which may bypass rate-limiting steps present in the conventional Δ6-desaturase/elongase pathways. The accumulation of EPA and AA in transgenic plants is a breakthrough in the search for alternative sustainable sources of fish oils.&nbsp