Cloning and bioinformatic characterization of genes controlling key steps of the fatty acid biosynthesis and lipid breakdown in seeds of Jatropha curcas L

Jatropha curcas L. (2n=2x=22) is becoming a popular non-food oleaginous crop in several developed countries for its proposed value in the biopharmaceutical industry. Despite the potentials of its oil-rich seeds as a renewable source of biodiesel and an interest in large-scale cultivation, relatively little is known with respect to population genetics and plant genomics. We recently performed genomic DNA markers and FCSS analyses to gain insights on ploidy variation and heterozygosity levels of multiple accessions, and genomic relationships among commercial varieties and locally dominant ecotypes grown in different geographical areas. Seeds commercialized worldwide seem to include a few closely related genotypes showing high degrees of homozygosity for single varieties and very low genetic diversity between varieties. For a better understanding of oil production and accumulation in J. curcas seeds, it would be useful to clone and characterize genes controlling key steps of FA biosynthesis and lipid breakdown pathways. Gene bank searches and bioinformatic analyses allowed us to select 13 genes encoding for enzymes involved in lipid metabolism. Five of these genes were previously isolated in J. curcas (i.e., acetyl-CoA carboxylase I, 3-ketoacyl-ACP I and III, acyl-ACP desaturase, and acyl-ACP thioesterase II), and their nucleotide and amino acid sequences are available in the NCBI databases. The remaining eight genes were cloned in the present study (i.e., ATP-citrate lyase, acetyl-CoA carboxylase II, 3-ketoacyl-ACP II, 3-ketoacyl-ACP reductase, 3-hydroxyacyl-ACP dehydrase, enoyl-ACP reductase, acyl-ACP thioesterase I, and acyl-CoA dehydrogenase). Replicated cDNA samples were produced by retrotranscription of mRNAs isolated from mature seeds belonging to different commercial varieties and Mexican ecotypes. Gene expression levels were assayed by quantitative Real-Time PCR with gene-specific primer combinations. The haplotyping of single accessions will be attempted using a worldwide collection of J. curcas materials to find out the most representative and discriminant SNPs related to FA biosynthesis and lipid metabolism genes.