Genome-guided Molecular Characterization of Oil Genes in Coconut (Cocos nucifera L.)
Anand Noel C. Manohar1,3*, Darlon V. Lantican1,3, Melvin P. Dancel1,3,
Don Emanuel M. Cardona1,3, Alissa Carol M. Ibarra1, Cynthia R. Gulay1,3, Alma O. Canama1,
Roanne R. Gardoce1, and Hayde F. Galvez1,2
1Genetics Laboratory, Institute of Plant Breeding, College of Agriculture and Food Science,
University of the Philippines Los Baños, College, Laguna 4031 Philippines
2Institute of Crop Science, College of Agriculture and Food Science,
University of the Philippines Los Baños, College, Laguna 4031 Philippines
3Philippine Genome Center – Program for Agriculture, Livestock,
Fisheries and Forestry (PGC-Agriculture), University of the Philippines Los Baños,
College, Laguna 4031 Philippines
*Corresponding Author: This email address is being protected from spambots. You need JavaScript enabled to view it.
ABSTRACT
Coconut oil is a major source of medium chain fatty acids (MCFAs), which are health-promoting plant compounds. The MCFAs of coconut oil have been reported to exhibit various health properties such as antioxidant, antibacterial, antiviral, and cardiovascular benefits brought about by the multi-functionality of these complex MCFAs. Six (6) candidate genes involved in oil and MCFA synthesis were identified in the general seed oil biosynthetic pathway. The candidate gene sequences were mined using local BLAST in the coconut genome assembly constructed based on 15× PacBio® and 50× Illumina® MiSeq sequence reads of CATD coconut variety. Scaffolds harboring the candidate genes were mapped based on sequence homology alignment. Gene structures of all genes were elucidated using evidence-based and ab initio prediction algorithms. The coding DNA sequences of KasII and KasIII in coconut were characterized. These MCFA genes have not been characterized nor reported in coconut. Gene-specific PCR primers were designed targeting the coding regions of each gene. PCR conditions were optimized to mine natural allele variants across 48 established coconut varieties in the Philippines through EcoTILLING (Ecotype Targeting Induced Local Lesions IN Genomes). A single nucleotide polymorphism (SNP) on the lysophosphatidic acid acyltransferase genes (LPAAT) was detected in the ‘West African Tall’ (WAT) and ‘Aguinaldo Tall’ (AGDT) varieties. The partial LPAAT gene sequences of WAT and AGDT were cloned and sequenced in order to characterize the SNP. Based on the identified SNPs, robust DNA markers may be developed for high-throughput screening and selection of favorable alleles in genomics-assisted coconut breeding for outstanding high-quality oil producing varieties.
INTRODUCTION
The coconut (Cocos nucifera L.), dubbed as the “Tree of Life,” is the most important palm in the humid tropics, with approximately 12 million hectares planted with the crop in around 86 countries. In the Philippines, coconut is a major agricultural crop grown in around 3.6 million hectares of farmland or 26% of the country’s agricultural lands. The country produces an average of 13.8 million tonnes of harvest per year derived from 340 million bearing trees. As the second largest producer and exporter of coconut in the world, the industry provides employment to approximately 1.4 million Filipino coconut farmers who comprise 21.7% of the total number of farmers directly engaged in agriculture (FAO 2018). Every part of the coconut palm – from the leaves, all the way down to the roots – have various economic uses. Copra oil (or coconut oil), the coconut’s most economically important product, used to be the most-traded vegetable oil in the world (Purseglove 1985). However, there is a remarkable global decline in the importance of coconut oil when used for food based on consolidated values/reports from 1961 and 2011. One of the major reasons for this is the stigma associated with the presence of long-chain fatty acids (Nayar 2017). . . . read more
REFERENCES
BLACKBURN GL, kater g, masciolli ea, kowalchuk m, babayan vk, bistrian br. 1989a. a reevaluation of coconut oil’s effect on serum cholesterol and atherogenesis. J Philipp Med Assoc 65: 144–152.
Blackburn GL, Babayan VK. 1989b. Infant feeding formulas using coconut oil and the medium chain triglycerides. J Am Coll Nutr 8(3): 253–254.
Bach AC, Babayan VK. 1982. Medium-chain triglycerides: An update. Am J Clin Nutr 36(5): 950–962.
Barkley NA, Wang ML. 2008. Applications of TILLING and EcoTILLING as reverse genetic approaches to elucidate the function of genes in plants and animals. Curr Genomics 9(4): 212–226.
Bates PD, Stymne S, Ohlrogge J. 2013. Biochemical pathways in seed oil synthesis. Current Opinion in Plant Biology 16: 358–364.
Cantarel BL, Korf I, Robb SM, Parra G, Ross E, Moore B, Holt C, Alvarado AS, Yandell M. 2008. MAKER: An easy-to-use annotation pipeline designed for emerging model organism genomes. Genome Research 18(1): 188–196.
Cardona DEM, Manohar ANC, Isabedra ACI, Canama AO, Rivera RL, Reaño CE. 2015. SSR polymorphism analysis among coconut (Cocos nucifera L.) parental genotypes towards genetic mapping for copra oil yield and quality. Proceedings of the Federation of Crop Science Societies of the Philippines (FCSSP) 2015 Conference; Stotsenberg Hotel, Clark Freeport Zone, Pampanga, Philippines.
Chapman KD, Ohlrogge JB. 2012. Compartmentation of triacylglycerol accumulation in plants. J Biol Chem 297: 2288–2294.
Comai L, Young K, Till BJ, Reynolds SH, Greene EA, Codomo CA, Enns LC, Johnson JE, Burtner C, Odden AR, Henikoff H. 2004. Efficient discovery of DNA polymorphisms in natural populations by EcoTILLING. The Plant Journal 37(5): 778–786.
Cooper DN. 2010. Functional intronic polymorphisms: Buried treasure awaiting discovery within our genes. Hum Genomics 4(5): 284–288.
Dai X, Sinharoy S, Udvardi MK, Zhao PX. 2013. PlantTFcat: An online plant transcription factor and transcriptional regulator categorization and analysis tool. BMC Bioinformatics 14: 321.
Doyle JJ, Doyle JL. 1990. Isolation of plant DNA from fresh tissue. Focus 12: 13–15.
Dayrit FM, Buenafe OEM, Chainani ET, De Vera IMS. 2008. Analysis of monoglycerides, diglycerides, sterols, and free fatty acids in coconut (Cocos nucifera L.) oil by 31P NMR spectroscopy. J Agric Food Chem 56: 5765–5769.
[FAO] Food and Agriculture Organization. 2018. Coconuts, Production/Crops. Retrieved from http://faostat3.fao.org/browse/Q/Q/C/E on19 Feb 2018.
Fushiki T, Matsumoto K, Inoue K, Kawada T, Sugimoto E. 1995. Swimming endurance capacity of mice is increased by chronic consumption of medium-chain triglycerides. J Nutr 125(3): 531–539.
Galvez HF. 2016. Coconut genomics project 8 – Development of web-based breeding resource and Eco-TILLING towards insect resistance breeding in coconut. In: DOST Form No. 3B Semi-annual Progress Report (2015 Annual Technical Report). Laguna, Philippines: University of the Philippines Los Baños – Institute of Plant Breeding.
Kaunitz H, Slanetz CA, Johnson RE, Babayan VK, Garsky G. 1958. Nutritional properties of the triglycerides of medium chain-length. J Am Oil Chem Soc 35: 10–13.
Kintanar QL, Castro JS. 1988. Is coconut oil hypercholesterolemic and atherogenic? A focused review of the literature. Trans Nat Acad Science & Tech 10: 371–414.
Kozomara A, Griffiths-Jones S. 2014. miRBase: Annotating high confidence microRNAs using deep sequencing data. Nucleic Acids Research 42(D1): 68–73.
Knutzon DS, Hayes TR, Wyrick A, Xiong H, Davies HM, Voelker TA. 1999. Lysophosphatidic acid acyltransferase from coconut endosperm mediates the insertion of laurate at the sn-2 position of triacylglycerols in lauric rapeseed oil and can increase total laurate level. Plant Physiol 120: 739–746.
Korf I. 2004. Gene finding in novel genomes. BMC Bioinformatics 5: 59.
Lantican DV, STRICKLER SR, CANAMA AO, GARDOCE RR, MUELLER LA, GALVEZ HF. 2019. De novo genome sequence assembly of dwarf coconut (Cocos nucifera L. ‘Catigan Green Dwarf’) provides insights into genomic variation between coconut types and related palm species. G3: Genes, Genomes, Genetics 9: 6.
LIANG Y, YUAN Y, LIU T, MAO W, ZHENG Y, LI D. 2014. identification and computational annotation of genes differentially expressed in pulp development of Cocos nucifera L. by suppression subtractive hybridization. BMC Plant Biol 14:205.
Li-Beisson Y, Shorrosh B, Beisson F, Andersson M, Arondel V, Bates P, Baud S, Bird D, DEBONO A, DURRETT TP, FRANKE RB, GRAHAM IA, KATAYAMA K, KELLY AA, LARSON T, MARKHAM JE, MIGUEL M, MOLINA I, NISHIDA I, ROWLAND O, SAMUELS L, SCHMID KM, WADA H, WELTI R, XU C, ZALLOT R, OHLROGGE J. 2010. acyl-lipid metabolism. Arabidopsis Book 8: e0133.
Mensink RP, Zock PL, Kester AD, Katan MB. 2003. Effects of dietary fatty acids and carbohydrates on the ratio of serum total to HDL cholesterol and on serum lipids and apolipoproteins: A meta-analysis of 60 controlled trials. Am J Clin Nutr 77(5): 1146–1155.
Müller H, Lindman AS, Blomfeldt A, Seljeflot I, Pedersen JI. 2003. A diet rich in coconut oil reduces diurnal postprandial variations in circulating tissue plasminogen activator antigen and fasting lipoprotein (a) compared with a diet rich in unsaturated fat in women. J Nutr 133(11): 3422–3427.
Nayar NM. 2017. The Coconut: Phylogeny, origins and spread. Cambridge, MA: Academic Press. 240p.
Ogawa T, Uchimiya H, Kawai-Yamada M. 2007. Mutual regulation of Arabidopsis thaliana ethylene-responsive element binding protein and a floral homeotic gene, APETALA2. Ann Bot 99(2): 239–244.
Ohto MA, Fischer RL, Goldberg RB, Nakamura K, Harada JJ. 2005. Control of seed mass by APETALA2. Proc Natl Acad Sci USA 102(8): 3123–3128.
Orsasova J, Misurcova L, Ambrozova JV, Vicha R, Mlcek J. 2015. Fatty acids composition of vegetable oils and its contribution to dietary energy intake and dependence of cardiovascular mortality on dietary intake of fatty acids. Int J Mol Sci 16(6): 12871–12890.
Padolina WG, Lucas LZ, Torres LG. 1987. Chemical and physical properties of coconut oil. Philipp J Coconut Studies 12: 4–17.
Purseglove JW. 1985. Tropical Crops: Monocotyledones. 5th ed. Longman, London and New York. p. 440–450.
Scalfi L, Coltorti A, Contaldo F. 1991. Postprandial thermogenesis in lean and obese subjects after meals supplemented with medium-chain and long-chain triglycerides. Am J Clin Nutr 53(5): 1130–1133.
REYNOLDS KB, CULLERNE DP, EL TAHCHY A, ROLLAND V, BLANCHARD CL, WOOD CC, SINGH SP, PETRIE JR. 2019. Identification of genes involved in lipid biosynthesis through de novo transcriptome assembly from Cocos nucifera developing endosperm. Plant and Cell Physiology 60(5): 945–960.
Singh R, Ong-Abdullah M, Low ETL, Manaf MAA, Rosli R, Nookiah R, Ooi LCL, Ooi SE, Chan KL, Halim MA, Azizi N. 2013. Oil palm genome sequence reveals divergence of interfertile species in Old and New worlds. Nature 500(7462): 335–339.
Wang N, Shi L, Tian F, Ning H, Wu X, Long Y, Meng J. 2010. Assessment of FAE1 polymorphisms in three Brassica species using EcoTILLING and their association with differences in seed erucic acid contents. BMC Plant Biology 10:137–148.
XIAO Y, XU P, FAN H, BAUDOUIN L, XIA W, BOCS S, XU J, LI Q, GUO A, ZHOU L, WU Y, MA Z, ARMERO A, ISSALI AE, LIU N, PENG M, YANG Y. 2017. The genome draft of coconut (Cocos nucifera L.). Gigascience 6(11): 1–11
