Wansheng Chen

Depart of Pharmacy, Changzheng Hospital, Second Military Medical University, shanghai, China

Quality is the basis for the efficacy of Traditional Chinese Medicine (TCM), affecting herbs, formulations, and even the practice of TCM itself. In our laboratory, we used Isatis indigotica, a prevalent Chinese medicinal herb, as a model to illustrate strategies and methods for TCM Quality-design study.

First of all, tetraploid I. indigotica (2n=28) with better yield, higher antiviral activity and enhanced resistance was obtained from its natural diploid progenitor (2n = 14) after selection for five years. Chemical investigation demonstrated the lignans including lariciresinol and larch lignan glycosides represented important efficacious substances for the antiviral effect, which accumulated more in tetraploid I. indigotica than diploid progenitor. A further comprehensive survey of global gene expression performed by using an Arabidopsis thaliana whole genome Affymetrix gene chip revealed the variation of gene expression between autotetraploid and diploid I. indigotica, providing a pool of candidate genes for improving I. indigotica quality through transgenic manipulation.

Next, in order to facilitate the process of isolating these quality-related genes (especially gene family) such as that involved in biosynthesis of effective metabolites, I. indigotica transcriptome sequencing was performed. By using this gene database, a large number of quality-related genes were isolated and intensively investigated, including stress resistance-involved transcription factor family IiWRKYs, signal transduction-involved gene IiCPK1 and IiCPK2, plant development-involved gene IiSDD1, lignans biosynthetic pathway regulatory factor family IibHLHs, and a series of lariciresinol biosynthetic pathway genes such as IiPAL (DQ115905), IiC4H (GU014562), Ii4CL (GU937875), IiCCR (GQ872418), IiCAD (GU937874), IiC3H (JF826963), IiCCoAOMT (DQ115904), as well as IiDirs and IiPLRs gene families.

Lastly, gene-based metabolic engineering study was performed to improve quality. For example, overexpression of IiPLR1 greatly enhanced lariciresinol production in I. indigotica hairy root cultures with the content of 353.9 μg/g of dry weight, which was appropriately 6.3-fold more than that in the wild-type counterpart; transgenic I. indigotica expressing Bt Cry1Ac and Pinellia ternata agglutinin gene (pta) significantly enhanced plant tolerance to moths and aphids.

In conclusion, our study not only prompted the possibility to obtain I. indigotica with higher quality, but also provided a reference for the study of TCM Quality-Design.