| Allium wallichii Kunth is a perennial herb that has both ornamental and medicinal value.Wild Allium wallichii Kunth which is dominant species in this distribution range has become an important resource for tourism development in Hezhang,Bijie,Guizhou.However,with the urbanization development in recent years,the growing environment of Allium wallichii Kunth has been threatened,resulting in a large reduction of wild resource.Therefore,it is important to enhance the conservation of wild Allium wallichii Kunth.In this study,we investigated the physiological and biochemical changes in the process of dormancy of Allium wallichii Kunth seeds,identified chromosome ploidy,estimated genome size and analyzed the molecular mechanism of dormancy release of Allium wallichii Kunth seeds under suitable temperature conditions(25℃)using transcriptomic methods,for establishing a suitable method for germination of Allium wallichii Kunth seeds and providing scientific basis for the discovery and conservation of wild Allium wallichii Kunth resources.The main results are as follows:1.The results showed that the tested Allium wallichii Kunth was tetraploid with a genome size of about 33.02 Gb;the chromosome number of tetraploid Allium wallichii Kunth was 4n=4x=28,and the flow cytometry test was consistent with the identification results of root tip press method.2.The composition of wild Allium wallichii Kunth seeds and their differences from cultivated Allium tuberosum Rottl seeds were analyzed.A total of 782 metabolites were detected by UPLC-MS/MS analysis of Allium wallichii Kunth seeds and cultivated Allium tuberosum Rottl seeds via widely targeted strategy.The top 20 metabolites with up-and down-regulation included amino acids and their derivatives,triterpenoid saponins,steroid saponins,flavonols,flavones,isoflavones,free fatty acids,phenolic acids,alkaloids,and indole alkaloids.A total of 84 metabolic pathways with differences were annotated in the KEGG Pathway,and four which were significantly enriched differential metabolite pathways(p<0.1)including flavonoid and flavonol biosynthesis,tropane,piperidine and pyridine alkaloid synthesis,flavonoid biosynthesis and phenylpropanoid biosynthesis.3.The cytological characteristics and dormancy properties of the seeds of Allium wallichii Kunth were investigated.The results shows that the seeds were composed of seed coat,embryo and endosperm,and the cotyledons were not obvious.The extracts from the seeds of Allium wallichii Kunth significantly reduced the germination rate of rapeseeds and inhibited the elongation of the radicles of rapeseeds,which confirmed the existence of endogenous germination inhibitory substances.With the dilution of the extract,this inhibitory effect would be weakened,and the inhibitory effects of different polar extraction solvents were Methanol phase > Ethyl acetate phase > Acetone phase > N-hexane phase >Water phase,with a wide range of inhibitor polarity distribution.Combined with the results of metabolomics analysis,organic acids,phenolic acids,aldehydes,unsaturated lactones and phenolic compounds,alkaloids and other substances that inhibit seed germination were present in the seeds of Allium wallichii Kunth.4.In this study,four different temperatures(15℃,20℃,25℃ and 30℃)were used to treat the seeds of Allium wallichii Kunth.The results showed that the temperature significantly affected the germination of Allium wallichii Kunth seeds,and 25℃ was the optimum germination temperature.Beyond 25℃,Allium wallichii Kunth had the highest germination rate of 84.3%,which was significantly higher than that of 52.9% at 15℃,71.6%at 20℃ and 75.4% at 30℃.The results of physiological and biochemical indicators showed that the starch content decreased the most under 25℃ treatments during the germination process,the soluble sugar content before dewetting was significantly higher under 25℃treatments than 15℃ treatment,and the peroxidase and 6-phosphoglucose dehydrogenase activities were higher under 25℃ treatment than 15℃ and 20℃ treatment.5.By performing RNA-seq analysis on the seeds of Allium wallichii Kunth at three different stages of imbibition(PD),dewlap(DR)and germination(ND)under 15℃ and25℃ treatments.Analysis of differentially expressed unigene revealed that temperature affects the activation and repair of cells,catabolic utilization of storage materials,energy metabolism and other processes through affecting DNA replication,mismatch gene repair,and cell signal transport,which regulates germination of Allium wallichii Kunth seeds.In this study,the seeds of Allium wallichii Kunth were taken as the research object,and the chromosomal ploidy,size of nuclear genome,composition and germination mechanism of dormancy release of Allium wallichii Kunth were systematically studied. |
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