Global Transcriptional Profiling Of The Developing Seeds For Characterizing Molecular Basis Of Oil Accumulation In Millettia Pinnata

Pongamia(Millettia pinnata)is an oilseed legume species which has high yield of nonedible seed oils and emerging as potential feedstock for sustainable biodiesel production.Breeding Pongamia for favorable traits in commercial application will rely on a comprehensive understanding of molecular mechanism regulating oil accumulation during its seed development.To date,only limited genomic or transcript sequences are available for Pongamia,while a temporal transcriptome profiling of developing seeds is still lacking in this species.We performed high-throughput sequencing for the representative RNA samples from three developmental phases of legume seeds and generated a dataset providing a panoramic view of gene expression dynamics during seed development.In this work,we conducted a time series analysis of morphological and physiological characters,oil contents and compositions,as well as global gene expression profiles in developing Pongamia seeds.Firstly,the three major developmental phases were characterized based on the combined evidences from embryonic shape,seed weight,seed moisture content,and seed color.Then,the gene expression levels in these three phases were quantified by RNA-seq analyses with three biological replicates from each phase.A high proportion(93.88%)of unigenes were expressed at all three phases,whereas less than 2% of unigenes were exclusively expressed at one of the three phases.A total of 8,881 differentially expressed genes(DEGs)were identified in at least one comparison between phases.Furthermore,the qRT-PCR analyses for 10 candidate DEGs involved in lipid metabolism demonstrated a good reliability of our RNAseq data in temporal gene expression profiling.We observed a dramatic increase in seed oil content from the embryogenesis phase to the seed-filling phase,followed by a steady and moderate increase towards the maximum at the desiccation phase.Seven types of fatty acids,including palmitic acid,stearic acid,oleic acid,linoleic acid,linolenic acid,eicosanoic acid,and behenic acid,were detected in all samples of seed oils.the relative proportion of oleic acid was 43.33%at 30 WAF,and became the most abundant type of fatty acid in Pongamia seed oils since 22 WAF.We proposed that a highly active expression of most genes related to fatty acid and triacylglycerol(TAG)biosynthesis at the embryogenesis phase might trigger both the substantial oil accumulation and the membrane lipid synthesis for rapid cell proliferation at this phase,while a concerted activation of TAG synthesis-related genes at the desiccation phase might further promote storage lipid synthesis to achieve the maximum content of seed oils.This study not only built a bridge between gene expression profiles and oil accumulation in developing seeds,but also laid a foundation for future attempts on genetic engineering of Pongamia varieties to acquire higher oil yield or improved oil properties for biofuel applications.