Cloning And Expression Analysis Of Chalcone Isomerase OfCHI From Osmanthus Fragrans

Osmanthus is a small evergreen tree species,is our country some species,has a long history.Nowadays,the research of Osmanthus fragrans is very extensive and deep,and has made remarkable achievements.In 2004,China has been granted the authority of the international registry of Osmanthus species,which is the second such authority in china.Osmanthus fragrans is rich in variety,and its varieties can be classified according to their different colors.Its color from shallow to deep mainly from milk white to orange red.The flower color of Osmanthus fragrans is one of the main bases for the classification of Osmanthus fragrans,and the study on the flower color of Osmanthus fragrans is of great significance in its classification and identification.It can also lay a theoretical foundation for the cultivation and breeding of new varieties of Osmanthus fragrans,promote the breeding of new varieties of Osmanthus fragrans,and promote the development of Osmanthus fragrans industry.The pigment of Osmanthus fragrans is mainly flavonoid,anthocyanin is a kind of flavonoids,which is the main material of flower color.At present,there are few reports about the flower color of Osmanthus fragrans,and the study of anthocyanin in Osmanthus fragrans is rare.To explore the effect of anthocyanin content of sweet scented osmanthus flower color,the expression of anthocyanin synthesis related enzyme CHI gene in anthocyanin biosynthesis and CHI gene in different tissues in osmanthus,based on the ‘Chenghongdangui',‘Jingui',‘Zaoyingui' 3 varieties as test osmanthus materials,determination of 3 varieties of sweet scented osmanthus anthocyanin content.At the same time,the use of RACE technology cloned the full-length cDNA of CHI gene of OfCHI,and use qRT-PCR analysis of the CHI gene in 3 kinds of sweet scented osmanthus and its expression in different tissues of characteristics.The main results are as follows:(1)‘Jingui' petals cDNA as template,CHI gene sequence in Osmanthus fragrans was cloned by technique of RACE.The sequence of CHI gene was 1069 bp,the coding region of 5' was 96 bp,and the coding region of 3' was 226 bp and have an open reading frame of 747 bp.Name this gene as OfCHI,GeneBank login number is KR815815.Sequence analysis showed that OfCHI amino acid sequence on the activation site of CHI protein: Arg-38,Gly-39,Leu-40,Phe-49,Thr-50,Tyr-108,Lys-111,Val-112,Asn-115,and Cys-116.(2)At the same time,used RT-PCR analysis of expression characteristic of OfCHIin 3 kinds of osmanthus fragrans.To analyze the expression characteristics of different varieties of sweet scented osmanthus,the expression of OfCHI in the‘Chenghongdangui' is the highest,in the ‘Zaoyingui' and ‘Jingui' is low.The expression of the gene in Osmanthus fragrans was positively correlated with anthocyanin content.When analyzing the expression of gene in different organs of Osmanthus fragrans,The expression of OfCHI in 3 species of Osmanthus fragrans was the highest in leaves,and was much higher than that in the stems and flowers,in the middle of the stem,the lowest in the flower.The results showed that in the 3 kinds of sweet scented osmanthus,the difference of the expression of leaf and flower and stem was significant,but the difference was not significant.(3)The anthocyanin content in petals of different cultivars of Osmanthus fragrans was tested.The anthocyanin content in ‘Chenghongdangui' is highest,which is 1.57mg/100 g,when the anthocyanin content of ‘Jingui' and ‘Zaoyingui' are low,respectively: 0.19mg/100 g and 0.06 mg/100 g.The anthocyanin content is differences significant in ‘Chenghongdangui' with ‘Jingui' and ‘Zaoyingui',while ‘Jingui' and‘Zaoyingui' between the difference was not significant.