Identification And Computational Annotation Of Genes Differentially Expressed In Pulp Development Of Cocos Nucifera L.

Coconut (Cocos nucifera L.) is one of the world’s most versatile, economically important tropical crops. Little is known about the physiological and molecular basis of coconut pulp (endosperm) development and only a few coconut genes and gene product sequences are available in public databases. This study identified genes that were differentially expressed during development of coconut pulp and functionally annotated the identified genes using bioinformatics analysis.Pulp from three different coconut developmental stages was collected. Suppression subtractive hybridization (SSH) libraries were constructed, and identified sequences were computationally annotated using Blast2GO software. A total of1272clones were obtained for analysis from four SSH libraries with63%showing similarity to known proteins. Pair-wise comparing stage-specific gene ontology (GO) ids from libraries B-D, A-C, B-C and A-D,6genes were continuously upregulated and28genes were downregulated;28genes were transiently upregulated and9genes were downregulated. Using KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis,1-acyl-sn-glycerol-3-phosphate acyltransferase (LPAAT), phospholipase D, acetyl-CoA carboxylase carboxyltransferase beta subunit,3-hydroxyisobutyryl-CoA hydrolase-like and pyruvate dehydrogenase E1β subunit were associated with fatty acid biosynthesis or metabolism. Triose phosphate isomerase, cellulose synthase and glucan1,3-β-glucosidase were related to carbohydrate metabolism, and phosphoenolpyruvate carboxylase (PEPC) was related to both fatty acid and carbohydrate metabolism. Of737unigenes,103encoded enzymes were involved in fatty acid and carbohydrate biosynthesis and metabolism, and a number of transcription factors and other interesting genes with stage-specific expression were confirmed by real-time PCR (RT-PCR), with validation of the SSH results as high as66.6%. Based on the determination of coconut endosperm fatty acids content by gas chromatography-mass spectrometry, a number of candidate genes in fatty acid anabolism were selected for further study.Functional annotation of genes differentially expressed in the coconut pulp development helped us understand the intrinsic link between coconut endosperm development and its molecular basis. By the SSH method, we obtained many genes related to fatty acid, carbohydrate and secondary metabolites, some transcription factors genes and several stress response genes, the results will have a great importance in understanding gene functions and regulatory networks in coconut fruit.