Effects Of Short-day Treatment On Expression Of Co Gene And Content Of Linarin In Chrysanthemum Indicum L.

The flower of Chrysanthemum indicum L.is the dry capitulum of Chrysanthemum indicum L.,which has the functions of clearing heat,detoxifying,purging fire and calming liver.The main chemical constituents of C.indicum include flavonoids,phenolic acids,terpenes and volatile oil.It has many pharmacological functions,such as anti-inflammatory,antibacterial,anti-oxidation,anti-virus,anti-tumor,liver protection and so on.In recent years,with the development and utilization of C.indicum resources,the market demand for raw materials of C.indicum is increasing.C.indicum is a typical short-day plant.Its biological characteristics of autumn flowering can not meet people's annual demand for its products.In recent years,scholars have studied a variety of ways related to regulating plant flowering,and gradually found out the gene network related to plant flowering.In the study of the photoperiod pathway of plant flowering,CO is found to be the key gene in the photoperiod flowering pathway of model plant Arabidopsis.Therefore,based on transcriptome sequencing analysis of C.indicum,this study simulated the short-day environment needed for flower formation,and explored the effects of short-day treatment on the expression of CO and the accumulation of linarin,a key gene in the photoperiod pathway of C.indicum.The main research result obtained are as follows:(1)The transcriptome sequencing of C.indicum was performed using BGISEQ-500 platform of Huada Gene Company.6.66Gb data were measured.After assembly and de-redundancy,54112 Unigenes were obtained.The total length,average length,N50 and GC content were 4159 2321 bp,768 bp,1170 BP and 40.60%,respectively.The Unigene was compared with the seven major functional databases,and functional annotations were made.Finally,3881 Unigenes(NR:70.23%),24859(NT:45.94%),23827(SwissProt:44.03%),28556(KOG:52.77%),27437(KEGG:50.70%),12082(GO:22.33%)and 31601(InterPro:58.40%)were identified.Get function notes.31059 CDS were detected using Transdecoder.At the same time,5181 SSRs were detected in 4563 Unigenes and 1393 Unigenes encoding transcription factors were predicted.(2)The CDS sequence of the C.indicum CO gene was cloned by RT-PCR and analyzed by bioinformatics.The results showed that the open reading frame of the C.indicum CO gene was 1380 bp,encoding 459 amino acids,containing one B-BOX domain and one CCT domain.The molecular weight of the protein was about 52.04 kDa,the theoretical isoelectric point was 4.81,the ?-helix accounted for 17.65%,the random coil accounted for 76.69%,and the extension chain accounted for 5.66%.No ?-folding,no signal peptide.The results of phylogenetic tree analysis showed that the C.indicum CO had high homology with Chrysanthemum lavandulifolium and Artemisia annua,which showed that CO was more conservative with other plants in the same family.In order to study the effect of short-day treatment on CO expression of the key gene of photoperiod in C.indicum,Real-Time PCR was used to detect the relative expression of CO in different parts of the C.indicum and different days of short-day treatment.The results showed that CO expression in leaves of C.indicum was the highest and significantly different from that in other parts.The order of CO expression in different parts of C.indicum was leaves,stems,buds,flowers and roots.The CO gene expression of C.indicum in the control group and the test group was significantly different in different periods of short-day treatment.The CO expression in the test group was significantly lower than that in the control group within 18 days after short-day treatment,but increased significantly from 22 days later than that in the control group,and the gene expression increased most at 26 days.In order to investigate the effect of short-day treatment on the accumulation of active components of C.indicum,the C.indicum was treated with short-day treatment for one month,and the content of linarin in C.indicum was determined by high performance liquid chromatography.The results showed that the content of linarin in the control and test groups reached the linarin content of the pharmacopoeia.The content of linarin in the test group was 24.21 mg·g-1,which was 4.27 mg·g-1 lower than that of the control group.The short-day treatment would increase the accumulation of the linarin of C.indicum.