Cloning and expression pattern analysis of CoFPPS gene in Camellia oleifera seed
[Objective]For understanding the biological characteristics and functions of CoFPPS gene in Camellia oleifera,and the regulatory mechanism of CoFPPS gene in C.oleifera seed squalene synthesis pathway.[Method]C.oleifera germplasm'WBL'with high squalene content were used as experimental materials.Based on the RNA-seq,qRT-PCR and gas chromatography tandem mass spectrometry,FPPS genes were discovered from C.oleifera seed.Meanwhile,gene cloning,bioinformatics,gene expression and squalene content were analyzed,to understand the physicochemical properties of CoFPPS gene,and the relationship of expression pattern of CoFPPS gene and squalene content.[Result]The coefficient of homology similarity between CoFPPS gene and FPPS1 gene(XM_028249399.1,C.sinensis)was 99.13%in NCBI database,the open reading frame of CoFPPS was 1 029 bp,encoding 342 amino acids,no signal peptide,and located to the cytoplasm.The relative molecular mass of CoFPPS was 39 332.14 Da,which was acidic stable protein with no obvious transmembrane domain in the peptide chain.In addition,the secondary structure was mainly based on alpha helix(222,64.91%),the sequence similarity of protein tertiary structure was 82.94%,the value of QMEANDisCo Global was 0.81±0.05.The amino acid sequence of CoFPPS shared high homology with FPPS from Actinidia chinensis var.chinensis,Impatiens glandulifera,Pelargonium graveolens,Juglans regia L.,Arachis ipaensis and Glycine max,and they belong to the Isoprenoid-Biosyn-C1 superfamily.CoFPPS,ClFPPS,RgFPPS,RmFPPS,RsFPPS,VdFPPS,PnFPPS and PjFPPS were classified into the same branch.qRT-PCR results revealed that CoFPPS gene was significantly up-regulated expression during developing period in seed,and which was consistent with the dynamic change pattern of squalene contents.[Conclusion]CoFPPS is an isoprene synthetase,which may have similar biological functions with FPPS proteins of other species,and CoFPPS gene expression is associated with the biosynthesis of squalene in C.oleifera seeds.
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