| Flower color as a pivotal phenotype has been considered to the important basis to population diversity formation,underlying the distribution patterns of flower color variations within population and the molecular mechanism can provide a reference for analyzing the evolve of population color variations.Meanwhile,that can rich genetic resources for ornamental flower color breeding.In this study,a wild distribution flower color variations population,Pleione limprichtii,in Sichuan Huanglong Nature Reserve,was used as the research material.There are rose-purple,pink and white different color individuals,quantitatived the three distinct flower colors within population,described the variant types and quantity distribution pattern.UPLC flower pigments component analysis to determine anthocyanins and RNA-seq analysis was used to identify the cruical genes involved in flower color formation,induced the anthocyanin biosynthesis pathway of P.limprichtii,explaining the molecular mechanism of variations.The main results are as follow:(1)Artificial quantitative statistics of three distinct flower color individiuals distribute in three population of P.limprichitii,the results showed: The ratio of the number of plants,rose-purple flower: pink flower: white flower is roughly 6: 3: 1;RHCSS evaluation index indicates that the three colors plants all belongings purple group;CIEL*a*b* quantified color value parameter results indicated that the rose-purple flower has the highest chroma value(C*),while the white flower has the lowest chroma value,implying that it is possible to distinguish three different colors according to the chroma value in the transitional variant color population;according to the distribution pattern of the three flower colors in the population,we speculated that the pattern may be a reproduction strategy to maintain the max pollination efficiency in the population of P.limprichtii.(2)UPLC was performed to identify flower pigments,results showed that flower color formation of P.limprichtii involved Cyanidin branch and Delphindin branch.RNA-seq sequencing technology and de novo assembly of perianth and lips organs in three different flower colors.A total of 80,525 unigenes were obtained,21 structural genes that may be involved in flower color variations were identified and encoded 10 catalytic enzymes,primary mapped the anthocyanin biosynthesis pathway,and inferred that expression pattern divergence of Pl FLS,Pl ANS and Pl UFGT are crucial for color variation of P.limprichitii.(3)Analysis of transcriptome database,a total of 14 Unigenes homologous to R2R3-MYB in Arabidopsis thaliana,32 Unigenes homologous to b HLH and 23 Unigenes homologous to WD40 were selected,phylogenetic tree of 14 Pl MYB amino acid sequences of P.limprichiti and 121 R2R3-MYB family members in Arabidopsis identified that Pl MYB13 transcription factor in P.limprichiti may participate in the regulation of anthocyanin production,and Pl MYB10 may repress anthocyanin production.Co-expression regulatory network by constructing structural genes and transcription factors suggested that Plb HLH and Pl WD40 may interation with Pl MYB10 transcription factor to forms Pl MYB10/Plb HLH20/Pl WD40-1 complex or Pl MYB10/ Plb HLH26/Pl WD40-1 complex participation in the regulation of the expression pattern of Pl FLS in P.limprichiti,affecting the formation of flower color variation.To sum up,this study is the first to investigate the color distribution pattern of P.limprichti in the natural population,identify anthocyanin substance and regulated genes related to the formation of pigment synthesis.Conduct the Anthocyanin biosynthesis pathway.This study is of great significance for the collection and breeding of genetic resources of the Pleione genus,and provides a reference for the formation and evolution of orchid color and angiosperm variation in the future. |
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