Microscopic Observate To Haustorium Formation Process Of Phelipanche Aegyptiaca And Screen For Genes That Affects Its Formation

Objective:Phelipanche aegyptiaca is a root parasitic weed,which deriving nutrients and water from host plants using specialized feeding structures known as haustoria.Defining formation process of P.aegyptiaca haustorium and exploring formation mechanism of haustorium,maybe can provide effective theoretical support for the control of P.aegyptiaca.Method:We observated the process of haustorium formation through paraffin section and microscopic observation,detected the changes of endogenous hormones through liquid mass spectrometry（LC-MS）.Screening and analysising genes that related to the formation of P.aegyptiaca haustorium by transcriptome sequencing technology.Defined the formation mechanism of haustorium,and established tissue culture system of P.aegyptiaca and induced its callus parasite melonResult:1 On the 5th day of inoculation,P.aegyptiaca formed initial haustorium.On the 7th day,the haustorium top was firmly attached to the host root,and then haustorial cell invaded the cortex of melon root,which formed a large amount of claw-like structures in the cell layer of the cortex of melon root.On the 11th day,haustorial cells connected with host vascular bundle.2 Phytohormone test results showed that a total of 17 phytohormones was involved in P.aegyptiaca haustorium formation process.The contents of Auxins,Cytokinins and Brassinoids changed significantly.The fold difference in the content of Auxin indole-3-carboxaldehyde in the process of P.aegyptiaca formatted initial haustorium（S0 stage）to haustorial cells connected with host vascular bundle（S3 stage）was as high as 6.12.The content of Cytokinin-like trans-zeatin riboside increased about 6.02 times and Brassinolide increased by 481.95 times.We hypothesized that these three phytohormones played an important role in regulateing P.aegyptiaca haustrium formation.3 Transcriptome data analysis results showed a total of 71,258 differentially expressed genes were obtained,some of which were enriched in the biological process of endogenous plant hormone biosynthesis.What’s more,many genes were annotated in Auxin,Cytokinin,and Brassinolide biosynthesis synthetic pathway.Auxin synthesis-related genes had different gene expressions in each stage of haustorial formation,while Brassinolide-related genes were up-regulated expression at haustorial S0,S1,and S3 stages.The expression of Cytokinin synthesis-related genes was up-regulated in the S3 stage of the haustorium.It is speculated that the synthesis-related genes of Auxin,Brassinolide and Cytokinin play a crucial role in the formation of the P.aegyptiaca haustrium4 Using B5 medium without any phytohormones and B5 medium with 8 mg/L GA₃+1 mg/L IAA could efficiently induce P.aegyptiaca callus formation.The induction rates were 88.89%and 86.11%,respectively.The best medium for adventitious bud induction was B5+8 mg/L GA₃+1 mg/L IAA+1 mg/L6-BA,and the induction rate was 58.21%.Second,two-week P.aegyptiaca callus induced in B5 medium without any phytohormones could parasitize melon roots.Conclusion:Under root chamber conditions,11 days after inoculation,P.aegyptiaca haustoria matured.During the process of invading melon root cortex,haustorium cells form a large number of claw-like structures,which are conducive to their fixation and expansion;During the development of haustrium,the Auxin hormones,Cytokinins,Brassinosteroids and the genes controlling the synthesis of three types of hormones played important regulatory role.Secondly,using P.aegyptiaca seeds as explants can efficiently and stably induce callus and adventitious buds.