Preliminary Study On The Chromogenic Mechanism Of Leaf In Red-leaf Poplars

'Zhonghong'(Populus dehoides 'Zhonghong')and 'Quanhong'(Populus deltoids'quanhong'),as two new red-leaf poplar varieties have received considerable attention for its highly ornamental value.But the studies on the coloration mechanism of the red-leaf poplars are very scarce,which delay the further development and utilization of foliage plant with red-leaf poplars representative.Three basic kinds of pigments are found in the leaf of higher plants:chlorophyll,carotenoids and anthocyanins.The ratio of the three kinds of pigments and their distribution in the leaf determine the final colouration.In this study,to preliminarily investigate the physiological and molecular mechanism in red-leaf poplars,we discussed the synthetic changes of pigments,the expression of anthocyanin biosynthetic genes,the contents of seven precursors in the biosynthesis of chlorophyll,the expression of Chlorophyllase in the degradation of chlorophyll and the difference of chloroplast ultrastructure in 'Zhonghong','Quanhong' and Populus Linn 2025,The main results were showed as follow.1.The analysis of different pigments accumulation in three poplarsCompared to Populus Linn 2025,a significantly higher accumulation of anthocyanin and a lower accumulation of chlorophyll and carotenoid were detected in two red-leaf poplars.The ratio of anthocyanin and total chlorophyll was also higher in red-leaf poplars than in the green one.These results suggested that the coloration of red leaves in red-leaf poplars was the result of anthocyanin degradation and chlorophyll synthesis.2.Cloning and expression analysis of anthocyanin biosynthetic genesAccording to the consensus sequences of anthocyanin synthesis genes in Populus trichocarpa,specific primers were used to amplify the six structural genes from Populus Linn 2025 and the sequence data was submitted to GenBank.Partial sequences were obtained for the following genes:CHS(GenBank Accession:JQ065897),CHI(GenBank Accession:JQ315243),F3H(GenBank Accession:JQ315246),F3 'H(GenBank Accession:JQ315245),F3'5'H(GenBank Accession:JQ315244),DFR(GenBank Accession:JQ065898),ANS(GenBank Accession:JQ065896)and UFGT(GenBank Accession:JQ315247).A real-time PCR analysis showed that the expression of all eight structural genes in the leaves of 'Quanhong' was much higher than in the green one,and enzymes F3 'H,DFR and ANS,which showed highest expression levels,were likely to be the key enzymes regulating the accumulation of anthocyanin in'Quanhong'.However,the same genes showed no detectable difference in gene expression between 'Zhonghong' and Populus Linn 2025 and only genes F3H,F3 'H,DFR and ANS expressed higher than the control.3.Analysis of chlorophyll metabolic pathwayThe chlorophyll in higher plants was in dynamic changes of synthesis and degradation.Seven kinds of precursors in the biosynthesis of chlorophyll in leaf of three populars were measured.The results showed that there were no significant difference in the contents of AlA,PBG,Urogen?,Coprogen ? between red-leaf populars and the green one,but the contents of Proto?,Mg-Proto? and Pchlide in two red-leaf populars were lower or equal than in that of green one.It indicated that the synthesis of chlorophyll was inhibited during these steps,and this might led to the lower chlorophyll contents and red leaf phenotype.Chlorophyllase is the rate-limiting enzyme in the degradation of chlorophyll.The gene expression level of chlorophyllase analysized by real-time quantitative PCR showed 2.7 times higher in 'Quanhong' but 1/3 times lower in 'Zhonghong' than in that of Populus Linn 2025.These results indicated that the increased gene expression level of chlorophyllase in the leaf of 'Quanhong' resulted in the speed up chlorophyll degradation and red leaf color of 'Quanhong'.But whether the reduction of chlorophyll content in'Zhonghong' was due to the degradation of chlorophyll still needs further research.4.The change of chloroplast ultrastructreIn this research,red-leaf poplars and the green one were quite different in chloroplast membrane structures.The most significant differences were the grana lamella became less and loose,the starch grain decreased and the osmiophilic globules increased.These results indicated that a decline in chloroplasts function could affect the synthesis of chlorophyll in red-leaf poplars.