| Gerbera jamesonii Bolus is a perennial herb of Compositae and one of the most popular cut flowers in the world.In this paper,the object of study is G.jamesonii`Jinxiang’,a single-petal golden variety with beautiful flower pattern and bright golden yellow color.The changes of carotenoid content in flower color were determined by Exogenous Gibberellin（GA3）and its inhibitor（PAC）.Real-time fluorescence quantitative PCR,hi TAIL-PCR,yeast single hybridization,double luciferase and VIGS were used to study the regulation of MYB on carotenoid synthesis and its response to carotenoid metabolism in the tongue flower of G.jamesonii`Jinxiang’.This study will lay a foundation for exploring the regulatory network of flower color formation.The main results are as follows:1.After GA3treatment,the color of the tongue-shaped flower of G jamesonii`Jinxiang’changed from golden to orange,and the carotenoid content increased by74.1%compared with the control at blooming stage.The expression of Gh PSY,a key gene in carotenoid biosynthesis pathway,increased by 53.1%after GA3treatment.The relative expression of Gh LYCB increased after GA3 treatment,while Gh LYCE did not change much.2.After GA3and PAC treatment,the expression levels of transcription factors Gh MYB1,Gh MYB2,Gh MYB3,Gh MYB6,Gh MYB7 and Gh MYB8 were consistent with Gh PSY.In addition,Gh MYB1,Gh MYB2,Gh MYB3,Gh MYB7,Gh MYB8transcription factors were consistent with the expression pattern of Gh PSY genes in different floral development stages.3.Eight full-length sequences of R2R3-MYB transcription factors were cloned from the tongue-shaped flower of G.jamesonii’Jinxiang’,were named as Gh MYB1,Gh MYB2,Gh MYB3,Gh MYB4,Gh MYB5,Gh MYB5,Gh MYB6,Gh MYB6,Gh MYB7,Gh MYB8,respectively.Its ORF length was 1093 bp,521 bp,534 bp,1125 bp,1331bp,540 bp,669 bp and 744 bp,encoding 364,174,180,375,443,180,222 and 247amino acids,respectively.Amino acid sequence analysis showed that Gh MYB3,Gh MYB7 and Gh MYB8 all belonged to R2R3-MYB transcription factor.The layout of hierarchical structure has certain similarities,which may indicate that their formal functions are also similar.4.The upstream sequence of Gh PSY 5’end was cloned by hi TAIL-PCR.The results of yeast single hybridization analysis showed that Gh PSY promoter could bind to Gh MYB2,Gh MYB7 and Gh MYB8 in yeast.Double luciferase assay showed that Gh PSY promoter could bind to Gh MYB2,Gh MYB7 and Gh MYB8.The activity was3.3,3 and 2 times higher than CK respectively.5.By agrobacterium tumefaciens the Gh PSY and Gh MYB3,Gh MYB7 and Gh MYB8.The results showed that,compared with CK,after instantaneous silencing of Gh PSY,Gh MYB3 and Gh MYB7,the content of carotenoids was decreased by 27.1%,61.8%and 30.5%respectively.After silencing Gh MYB3 and Gh MYB7,Gh PSY decreased by 4.7 and 5.7 times,respectively.Indicating that virus silencing of Gh PSY,Gh MYB3 and Gh MYB7 could reduce the synthesis of carotenoids,and that virus silencing of Gh MYB3 could not be achieved in some lingual flower roots. |
