Morphology And Transcriptomics Study Of Seed Maturity Of Panax Notoginseng

Sanqi（Panax notoginseng（Burk.）F.H.Chen）is a valuable traditional Chinese herbal medicine with³00 years history.Seed propagation is the main method for reproduction.The seed is incompletely matured when it has been harvested since it has features of after-ripening.It harms the production,because the seed life is short（15days）under natural conditions.Non-germinated state can keep for 40⁶0 days by using the most appropriate preservation method which is room temperature stratification.In this study,by morphological observation and transcriptome analysis,we analyzed the changes of seed in morphology and gene expression at the after-ripening stage.Our work provided a better understanding of seed germination,and presented a theoretical reference for regulation of germination time of sanqi seeds.The main results are as follows:1.Our data of germination rate and mildew rate supported that room temperature stratification is crucial for seed preservation.After being laminated for 40 days,seed started to germinate.After 40⁵0 days,when seed germination and mildew peaked in this period,seeds required ventilation.To preserve the seed vigor and to reduce seed deterioration,the wet sand should be kept in appropriate humidity and temperature.2.Through anatomic observation of the seed and embryo of Panax notoginseng,we observed the process of seed germination and the tendency of leaf primordium continuous differentiation during the stratification,which provided valuable referenced information for transcriptome studies in sampling and analyzing.3.Four key time nodes（（1）,（3）,（4）,（6））in anatomical observation were selected for transcriptome analysis,corresponding to A（seeds with embryos quarter of the seed length and peep plumule）,B（seeds with embryos growing to three quarters of the seed length and the radicle not yet breaking through endosperm）,C（the seeds of this period with the apex of the radicle breaks through the endosperm and its extra-seed length grows to 1/8 of the seed diameter）,D（seeds with external radicle length about half of the seed diameter）.We obtain embryo and endosperm of seed at four critical time nodes,transcriptome sequencing use Illumina HiSeq?4000 platform and assemble by Trinity,finally produced 351,327 transcripts and 117,227 unigenes,and N50 is 1,821 bp and 1,467 bp,respectively.4.Assaying and analysis of plant endogenous hormone in embryo showed that the concentration of Gibberellic acid（GA₃）played an important role in the period from A to B.At this stage,seed radicle was protected by the endosperm,which was the golden period to control the germination time of seeds.5.Through calculating the FPKM of candidate genes in embryo and endosperm,we found that the expressions of SUS gene was particularly prominent in B phase.Stage B was the key node of embryo development from inside to outside.Thus,it is the best time to regulate the germination time by intervening auxin transport and sucrose utilization.Therefore,we suppose that GNOM and SUS genes provide references for regulating the germination time of sanqi seeds.6.Combined with differential expression gene analysis and KEGG annotation,we find that the specific differential expression genes in B and C phases are closely related to the opening of photosynthetic system.lhcb1 gene might provide gist for regulating germination time of Panax notoginseng seeds.