Tomato (Solanum lycopersicum) glandular trichomes function as biochemical factories that synthesize a diverse array of specialized metabolites. Terpenoids are the most diverse class of plant specialized metabolites, with volatile mono- and sesquiterpenes playing important roles in plant defense. Although the biosynthetic pathways of volatile terpenes in tomato glandular trichomes have been well described, little is known about their regulation. Here, we demonstrate that SlMYC1, a basic helix-loop-helix transcription factor, differentially regulates mono- and sesquiterpene biosynthesis in the type VI glandular trichomes of tomato leaves and stems. SlMYC1 functions as a positive regulator of monoterpene biosynthesis in both leaf and stem trichomes but as a negative regulator of sesquiterpene biosynthesis in stem trichomes. SlMYC1 is also essential for type VI glandular trichome development, as knocking down SlMYC1 led to the production of smaller type VI glandular trichomes at lower densities, and knocking out this gene led to their absence. Our findings reveal a role for SlMYC1 not only in type VI glandular trichome development but also in the regulation of terpene biosynthesis in tomato.

The CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins) system was first identified in bacteria and archaea and can degrade exogenous substrates. It was developed as a gene editing technology in 2013. Over the subsequent years, it has received extensive attention owing to its easy manipulation, high efficiency, and wide application in gene mutation and transcriptional regulation in mammals and plants. The process of CRISPR/Cas is optimized constantly and its application has also expanded dramatically. Therefore, CRISPR/Cas is considered a revolutionary technology in plant biology. Here, we introduce the mechanism of the type II CRISPR/Cas called CRISPR/Cas9, update its recent advances in various applications in plants, and discuss its future prospects to provide an argument for its use in the study of medicinal plants.

Background: Agarwood is an expensive resinous heartwood derived fromAquilariaplants that is widely used in traditional medicines, incense and perfume. Only wounded trees can produce agarwood, and the huge demand for the agarwood products has led allAquilariaspp. being endangered and listed in the Appendix II of the CITES (http://www.cites.org). The major components of agarwood are sesquiterpenes and phenylethyl chromones. Owing to a lack of genomic information, the molecular basis of wound-induced sesquiterpenes biosynthesis and agarwood formation remains unknown.Results: To identify the primary genes that maybe related to agarwood formation, we sequenced 2 cDNA libraries generated from healthy and woundedA. sinensis(Lour.) Gilg. A total of 89,137 unigenes with an average length of 678.65 bp were obtained, and they were annotated in detail at bioinformatics levels. Of those associated with agarwood formation, 30 putatively encoded enzymes in the sesquiterpene biosynthesis pathway, and a handful of transcription factors and protein kinases were related to wound signal transduction. Three full-length cDNAs of sesquiterpene synthases (ASS1-3) were cloned and expressed inEscherichia coli, and enzyme assays revealed that they are active enzymes, with the major products being δ-guaiene. A methyl jasmonate (MJ) induction experiment revealed that the expression ofASSwas significantly induced by MJ, and the production of sesquiterpenes was elevated accordingly. The expression of some transcription factors and protein kinases, especiallyMYB4,WRKY4,MPKK2andMAPK2, was also induced by MJ and coordinated withASSexpression, suggesting they maybe positive regulators ofASS.Conclusions: This study provides extensive transcriptome information forAquilariaspp.and valuable clues for elucidating the mechanism of wound-induced agarwood sesquiterpenes biosynthesis and their regulation.

Background: Bupleurum chinenseDC. is a widely used traditional Chinese medicinal plant. Saikosaponins are the major bioactive constituents ofB. chinense, but relatively little is known about saikosaponin biosynthesis. The 454 pyrosequencing technology provides a promising opportunity for finding novel genes that participate in plant metabolism. Consequently, this technology may help to identify the candidate genes involved in the saikosaponin biosynthetic pathway.Results: One-quarter of the 454 pyrosequencing runs produced a total of 195, 088 high-quality reads, with an average read length of 356 bases (NCBI SRA accession SRA039388). Ade novoassembly generated 24, 037 unique sequences (22, 748 contigs and 1, 289 singletons), 12, 649 (52.6%) of which were annotated against three public protein databases using a basic local alignment search tool (E-value ≤1e-10). All unique sequences were compared with NCBI expressed sequence tags (ESTs) (237) and encoding sequences (44) from theBupleurumgenus, and with a Sanger-sequenced EST dataset (3, 111). The 23, 173 (96.4%) unique sequences obtained in the present study represent novelBupleurumgenes. The ESTs of genes related to saikosaponin biosynthesis were found to encode known enzymes that catalyze the formation of the saikosaponin backbone; 246 cytochrome P450 (P450s) and 102 glycosyltransferases (GTs) unique sequences were also found in the 454 dataset. Full length cDNAs of 7P450s and 7 uridine diphosphateGTs (UGTs) were verified by reverse transcriptase polymerase chain reaction or by cloning using 5' and/or 3' rapid amplification of cDNA ends. TwoP450s and threeUGTs were identified as the most likely candidates involved in saikosaponin biosynthesis. This finding was based on the coordinate up-regulation of their expression withβ-ASin methyl jasmonate-treated adventitious roots and on their similar expression patterns withβ-ASin variousB. chinensetissues.Conclusions: A collection of high-quality ESTs forB. chinenseobtained by 454 pyrosequencing is provided here for the first time. These data should aid further research on the functional genomics ofB. chinenseand otherBupleurumspecies. The candidate genes for enzymes involved in saikosaponin biosynthesis, especially theP450s andUGTs, that were revealed provide a substantial foundation for follow-up research on the metabolism and regulation of the saikosaponins.

Only when Aquilaria spp. or Gyrinops spp. trees are wounded, due to insect attack, or microbial invasion, agarwood can be successfully induced. In the present study, a fungus which can induce agarwood formation efficiently was isolated and a suitable method for its application to induce agarwood formation was developed. Rigidoporus vinctus was isolated from the inner layers from infectious A. sinensis trees. When the fermentation liquid of fungi inoculated back to A. sinensis tree, agarwood was found to be induced. In addition, a novel method called trunk surface agarwood-inducing technique (Agar-Sit) was developed to produce agarwood with R. vinctus. The alcohol soluble extract content of the agarwood, up to 38.9%, far higher than the requirement (10%) in Chinese Pharmacopoeia and the six characteristic compounds of agarwood used as Chinese Medicinal Materials were all detected. Their relative percentages of the sesquiterpenes in the essential oil were 22.76%. This is the first report of the Agar-Sit and also the application of R. vinctus in agarwood induction. According to the results, when the combination of Agar-Sit and R. vinctus is used agarwood can be induced with high yield and good quality.

Background: Chaihu is a popular traditional Chinese medicine that has been used for centuries. It is traditionally used to treat cold fever and liver-related diseases. Saikosaponins (SSs) are one of the main active components of chaihu, in addition to essential oils, flavonoids, and polysaccharides. Considerable effort is needed to reveal the biosynthesis and regulation of SSs on the basis of current progress. Objective: The aim of this study is to provide a reference for further studies and arouse attention by summarizing the recent achievements of SS biosynthesis. Methods: All the data compiled and presented here were obtained from various online resources, such as PubMed Scopus and Baidu Scholar in Chinese, up to October 2019. Results: A few genes of the enzymes of SSs participating in the biosynthesis of SSs were isolated. Among these genes, only the P450 gene was verified to catalyze the SS skeleton β-amyrin synthase. Several UDP-glycosyltransferase genes were predicted to be involved in the biosynthesis of SSs. SSs could be largely biosynthesized in the phloem and then transported from the protoplasm, which is the biosynthetic site, to the vacuoles to avoid self-poisoning. As for the other secondary metabolites, the biosynthesis of SSs was strongly affected by environmental factors and the different species belonging to the genus of Bupleurum. Transcriptional regulation was studied at the molecular level. Conclusion: Profound discoveries in SSs may elucidate the mechanism of diverse the monomer formation of SSs and provide a reference for maintaining the stability of SS content in Radix Bupleuri.