Quality Evaluation Method Discussion Based On The The Functional Genes In Lonicera Japonica Thunb

Honeysuckle， as a kind of chinese commonly used drugs zhat has manypharmacological and health functions，is the dried buds or open flowers of Lonicerajaponica Thunb.． The2010edition of the People’s Republic of chinaPharmacopoeia take the content of chlorogenic acid and luteoloside as the mainindicators of the evaluation of honeysuckle’ quality．In this research，we analyzedthe function and structure of key enzyme genes in on the base of former researchresult of the honeysuckle transcriptome data，we filtered a gene that have intimatelyinterrelated with the occumulation of chlorogenic acid． We take this gene toprokaryotic expression，then make and filter antibody．Combined with enzymelinked immunosorbent assay(ELISA) and research of honeysuckle’ chemicalconstitution，we try to buiLD a mathematic model that can be used to discriminatethe quality of honeysuckle quickly and high throughput．In this research，results are as follows:(1) On the base of former research result of the honeysuckle transcriptome data，we get the key enzyme genes in honeysuckle active ingredient gene regulatorynetworks．(2) FatB which is the key enzyme gene in fatty acid metabolic pathways wascloned，and we analyzed it’s gene sequence and protein characteristics in Lonicerajaponica Thunb.、Lonicera japonica Thunb. var. chinensis (Wats.) Bak.、Lonicerahypoglauca Miq and Lonicera dasystyla Rehd. by bioinformatics，We speculate theFatB gene may has the same biological function with AtFatB．Gene expressionanalysis shows that there is no significantly difference of FatB orthologous attranscripted level in fresh buds between Lonicera japonica Thunb.、 Lonicerajaponica Thunb. var. chinensis (Wats.) Bak.、 Lonicera hypoglauca Miq andLonicera dasystyla Rehd.(3) We also cloned two key anzyme genes in isoprenoid metabolic pathway： IspE and IspH．Gene expression analysis shows that transcripted level of IspE andIspH in bud of Lonicera japonica Thunb. var. chinensis (Wats.) Bak. wassignificantly higher zhan in Lonicera japonica Thunb.，which will help for furtherreseach on the synthesis of terpenes，aroma and color．(4) In the biosynthetic pathway of chlorogenic acid and luteoloside，we got andcloned15key enzyme genes，and we analyzed its’ gene structure characteristics、gene expression pattern and relationships between gene expression and content ofchlorogenic acid and luteoloside by using bioinformatics、real-time PCR、enzymeactivity assay、HPLC and so on. We found the PA1gene may be the key gene thathas effect on content of chlorogenic acid.(5) By gene cloning and prokaryotic expression，we prepared and selectedpolyclonal antibody of PAL1．In the follow-up research，we will measure the protein content of PAL1inLonicera japonica Thunb.、Lonicera japonica Thunb. var. chinensis (Wats.) Bak.、Lonicera hypoglauca Miq and Lonicera dasystyla Rehd. by ELISA experiment．Byanalyzing the results of ELISA and HPLC, we try to buiLD a mathematic model thatcan quickly discriminate the quality of honeysuckle.