Cloning and Characterization of the Oleosin cDNA Isoforms from Coconut (Cocos nucifera L.)

Ellen  Regalado; Jorge Gil  Angeles; Abigail Joy  Rodelas; Roberta Garcia; Antonio Laurena; Evelyn Mae Tecson-Mendoza

Ellen Regalado Institute of Plant Breeding, Crop Science Cluster, College of Agriculture, University of the Philippines Los Baños, College, Laguna, Philippines
Jorge Gil Angeles Institute of Plant Breeding, Crop Science Cluster, College of Agriculture, University of the Philippines Los Baños, College, Laguna, Philippines
Abigail Joy Rodelas Institute of Plant Breeding, Crop Science Cluster, College of Agriculture, University of the Philippines Los Baños, College, Laguna, Philippines
Roberta Garcia Institute of Plant Breeding, Crop Science Cluster, College of Agriculture, University of the Philippines Los Baños, College, Laguna, Philippines
Antonio Laurena Institute of Plant Breeding, Crop Science Cluster, College of Agriculture, University of the Philippines Los Baños, College, Laguna, Philippines
Evelyn Mae Tecson-Mendoza Institute of Plant Breeding, Crop Science Cluster, College of Agriculture, University of the Philippines Los Baños, College, Laguna, Philippines

Keywords: oleosin, coconut, Cocos nucifera, isoforms, cDNA sequence, amino acid sequence

Abstract

Oleosins are proteins in the oil bodies of oil-bearing seeds. Two cDNAs of 500 and 300 bp, OLE500 and OLE300, were cloned by RT-PCR from coconut endosperm using a gene specific primer (FSPVLV) based on the conserved and characteristic proline knot of oleosins. Three distinct oleosins were shown by their cDNA sequences: OLE300a, OLE500a and OLE500c, with OLE500a being the major isoform. These isoforms were observed in solid endosperm of 6–7 and 9–10 month old samples but not in the mature 11–12 month sample. Southern blot analysis showed two copies of OLE 500a gene. The partial cDNA sequences of OLE300a, OLE 500a and OLE500c had coding regions of 176, 242, and 233 bp, which code for 58, 80 and 77 amino acids, corresponding to molecular weights of 6040, 8600 and 8566, respectively. Analysis of the amino acid sequences revealed the highly conserved hydrophobic domain from the right half of the proline knot, the C-terminal amphipathic region characteristic of oleosins and that the coconut oleosins are of the L-form due to the absence of the 18-amino acid insert characteristic of the H-form. OLE 500a had 77.0% similarity with OLE500c and 85.96% with OLE 300a and the latter had 91.23% similarity with OLE 500c. OLE 500a and OLE 500c exhibited highest similarity with oil palm oleosin (86%) and OLE300a with soybean oleosin (72%), respectively.