Genome-Guided EcoTILLING Approach Towards Improved Fatty Acid Biosynthesis in Coconut

Coconut 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 benefits such as antioxidant, antibacterial, antiviral, and cardiovascular brought about by the multi-functionality of these complex MCFAs; lauric acid being present in 40-50%. Advanced tools in genomics such as next-generation sequencing can aid in unravelling the molecular basis of oil biosynthesis in coconut. Six (6) candidate genes involved in the production of lauric acid and other MCFAs were identified in the general seed oil biosynthetic pathway. The candidate gene sequences were mined using local BLAST in a draft coconut genome assembly constructed from the 11X PacBio and 40X Illumina MiSeq sequence data of ‘Catigan Dwarf’ (CATD) coconut. Resulting contigs having significant BLAST hit were extracted prior to annotation. Gene structures of all candidate genes were predicted ab initio. Coding DNA sequences of KasII and KasIII in coconut were identified; whereas these genes are not yet reported previously through any conventional gene characterization approaches. Using the determined sequence information, gene-specific polymerase chain reaction (PCR) primers were designed. PCR conditions were optimized to mine the natural variants across 48 established coconut varieties in the Philippines via Targeting Induced Local Lesions In Genomes (EcoTILLING). SNP on LPAAT was detected on ‘West African Tall’ (WAT) and ‘Aguinaldo Tall’ (AGDT) varieties. SNPs were further characterized for high-throughput screening and selection of favorable alleles in genomics-assisted coconut breeding for outstanding high quality oil-producing varieties.