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Clone the Differential Expression Fragment from Ginseng Culture Induced by Salicylic Acid

Author: ZhangYue
Tutor: ZhangMeiPing
School: Jilin Agricultural University
Course: Crop biotechnology
Keywords: Panax ginseng C. A. Meyer elicitor Salicylic acid cDNA representational difference analysis
CLC: S567.51
Type: Master's thesis
Year: 2008
Downloads: 147
Quote: 3
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Abstract

The experiment presented in this paper is based on explants of two-year-old ginseng root, which are cell suspension cultured in medium consisting of MS+2,4-D(2.0mg/L)+BA (2.0mg/L)+IBA(5.0mg/L)+V_C(30mg/L).It is proved in the experiment that the content of ginsenoside can be improved 130%by adding salicylic acid in the logarithm stage of the ginseng cells compared with the reference,while it is not so effective if the salicylic acid is appended in the early stage of cell growth.It is also observed that the culturing stage can be extended by adding salicylic acid.Up to the 35th day in the ginseng callus tissue culturing,these with salicylic acid in it keep growing vigorously while those without salicylic acid begin declining.The content of peroxidase,polyphenol oxidase,phenylalanine ammonia-lyase is traced in 72 hours after the salicylic acid is appending.It is shown that the activity of peroxidase reached the peak after 24 hours,which is about 1.5 times of the reference.;the activity of polyphenol oxidase reaches the peak after 18 hours and remains more vigorous than the reference during 72 hours;the activity of phenylalanine ammonia-lyase rises fast after 24 hours and reaches the peak after 48 hours,which is about 1.87 times of the reference.The improvemenrt of activity of these three enzymes represents the enhancement of secondary metabolites,which The content of peroxidase,polyphenol oxidase,phenylalanine ammonia-lyase is traced in 72 hours after the salicylic acid is appending.It is shown that the activity of peroxidase reached the peak after 24 hours,which is about 1.5 times of the reference.;the activity of polyphenol oxidase reaches the peak after 18 hours and remains more vigorous than the reference during 72 hours;the activity of phenylalanine ammonia-lyase rises fast after 24 hours and reaches the peak after 48 hours, which is about 1.87 times of the reference.The impr6vemenrt of activity of these three enzymes represents the enhancement of secondary metabolites,which is beneficial to the synthesis and accumulation of secondary metabolites.Salicylic acid is a kind of very effective elicitor for a variety of medicinal plants used in cell culture,the effect significantly.However,the mechanism in which the salicylic acid induced the sythesis of plant secondary metabolites is not very clear till now.It is the first time that the cDNA-RDA technology is applied in the research of secondary metabolites of medicinal plants with the optimized technological system.It is determined that the total RNA should be abstracted after 24 hours when the acid is added.Before the different fragment is screened,the double-stranded cDNA should be synthesized by replacement and three pairs of jointing primer. a 516 bp fragment of the differences is obtained through the analysis of differential display.The homologous sequence of the gene is not found in the nucleotide sequence of GenBank,so the differential expression is considered to be part of a new gene that has not been separated.The registration number of the differential expression is FE900130 in GenBank.Because the former 217 bp fragment shows great homology with hexokinase,therefore it is speculated that the functions of the differential fragment should be similar with hexokinase at certain degree.More further work is called for to develop specific functions.

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