| Chrysanthemum vestitum[Chrysanthemum vestitum(Hemsl.)Ling]is closely related to modern chrysanthemum(Chrysanthemum X morifolium Ramat.).Both of them are hexaploid species of chrysanthemum genus.Their capitula are consisted of ray florets and disc florets.Privious investigators found some mutants whose ray florets seem like tubular petals in chrysanthemum.This variation attracts great attention of researchers.Thus,taking the mutant as material to investigate the mechanism behind the morphological variation could provide theoretical basis and guide for the breeding of chrysanthemum.In this work,variation of 14 main morphological characteristics in 10 C vestitum populations inhabiting three sites(Tianzhu mountain,Luanchuan,Neixiang)of Anhui and Henan province were investigated.Based on the analysis of morphological variation,RNA-seq and bioinformatics were conducted and analysised between two typical petal types.Meanwhile,differential expression genes and transcription factors were screened through FPKM.The qRT-PCR was taken to validate the result of RNA-seq.Through the analysis,several main results were obtained:(1)Through the analysis of 14 key morphological characteristics of C.vestitum in Anhui and Henan province,we found that the leaf width of C.vestitum in the two regions is almost the same,while the leaf of C.vestitum of Henan is much longer.This difference caused the disparity of leaf characteristic as the leaves of C.vestitum of Anhui is closer to ovate and the leaves of C.vestitum of Henan are longer and narrower.Analysis of characteristics among populations and regions indicate that the variation coefficient shows no significant difference in the same population to particular characteristics.Whithin the same population,leaf characteristics are more stable than the flower characteristics.We found that ray florets mutants of C.vestitum in Neixiang district of Henan province.According to their morphological characteristics,the mutants were divided into three types,flat type,tublar type and mix type.Through measurement,we found out that the basal fusion ratio of ray florets and the numbers of different petal types on one capitulum are also quantitative characters.(2)Through the RNA-seq of flat petals and tubular petals,92003 unigenes were obtained,and their average length are 861.06bp.NR,SWISSPROT,KOG,KEGG,GO databases were taken to provide annotation for the unigenes.And the total number of annotated unigene was 42381.Matching the unigenes to the Plant Transcription Factor Database v4.0,we identified 1232 unigenes,and they belong to 55 transcription factor families.66 of them belong to NAC family;39 belong to MADS-box family;19 belong to ARF family and 28 belong to TCP family.There were 160 differential expressed transcription factors between the two materials,12 of them belong to NAC transcription factor family;6 of them belong to MADS-box;3 of them belonged to TCP and ARF respectively.To validate the accuracy of RNA-seq,the author selected SAND as reference gene to calibrate the expression level of 12 genes selected from RNA-seq database.7 of them showed extremely significant correlation;4 of them showed significant correlation;only 1 gene didn't reach the significant level.To sum up,we drawed conclusions below,C.vestitum have different demands to different regions and they mainly distribute on the shady side of mountain top of Anhui Tianzhu mountain,and they distribute in thin forest of Henan region with the altitude limited to 400-1200m.The difference of leaf characters among populations and regions are greater,while the flower characters are less.The ray floret mutants were only found in Neixiang district,and the variation of ray florets was a quantitative character.Through the transcriptome analysis,160 differential expressed transcription factors were found,13 of them belong to NAC transcription factor family,6 of them belong to MADS-box,3 of them belong to TCP and ARF respectively.These genes may participat in the control of the formation of different petal types. |
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