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Background: Fuzi is a processed product of the lateral root of Aconitum carmichaelii Debx, a plant of the Ranunculaceae, and has been used to treat various diseases. This study used Illumina Hiseq High-throughput platform to sequence, assemble and annotate, screen development related genes, transcription pathways and functional enrichment in true root, lateral roots and “bridge”, and analyzed their correlations with the formation and development of lateral roots of A.carmichaelii, which can reveal the process and regulation mechanism of lateral roots growth and maturation of A. carmichaelii. Results: By sequencing, a total of 66.13Gb clean data and 28,982 unigenes with function annotation were finally obtained, with N50 of 1,627 bp, and 12,833 genes were assigned to 130 specific metabolic pathways by Kyoto Encyclopedia of Genes and Genomes (KEGG), then 2,599 were significantly differentially expressed. The KEGG analysis of the DEGs revealed that it was mainly enriched in starch and sucrose metabolism, ribosome, carbon metabolism, plant hormone signal transduction which play an important role in the expansion of lateral roots. The DEGs and pathways indicated that there was little differences between true root and “bridge”, while a big difference between them and lateral roots. The DEGs of auxin, cytokinin and other pathways may be conducive to the formation of lateral roots, which explained the development mechanism of lateral roots from a molecular point of view. Conclusions: This study provides a reference for the study of cultivation and management of lateral roots of A. carmichaelii.

Summary Magnolia officinalis, a representative tall aromatic tree of the Magnoliaceae family, is a medicinal plant that is widely used in diverse industries from medicine to cosmetics. We report a chromosome-scale draft genome of M. officinalis, in which ∼99.66% of the sequences were anchored onto 19 chromosomes with the scaffold N50 of 76.62 Mb. We found that a high proportion of repetitive sequences was a common feature of three Magnoliaceae with known genomic data. Magnoliids were a sister clade to eudicots-monocots, which provided more support for understanding the phylogenetic position among angiosperms. An ancient duplication event occurred in the genome of M. officinalis and was shared with Lauraceae. Based on RNA-seq analysis, we identified several key enzyme-coding gene families associated with the biosynthesis of lignans in the genome. The construction of the M. officinalis genome sequence will serve as a reference for further studies of Magnolia, as well as other Magnoliaceae., Graphical abstract, Highlights • We provide a chromosome-scale genome for Magnolia officinalis • We explore the phylogenetic position of Magnoliids • Analysis for lignan biosynthetic pathway is reported • We identify terpenes synthase in the floral fragrance composition of Magnolia, Biological sciences; Genetics; Evolutionary biology; Bioinformatics; Genomic analysis; Genomics

Magnolia officinalis, a representative tall aromatic tree of the Magnoliaceae family, is a widely used medicinal plants with diverse medicinal values and applications, such as afforestation, ornamental and so on. We report a chromosome‑scale draft genome of M. officinalis, in which ~ 99.66% of the sequences were anchored onto 19 chromosomes with the scaffold N50 of 76.62 Mb. We found that a high proportion of repetitive sequences was a common feature of three Magnoliaceae with known genomic data, and M. officinalis was the plant in Magnoliaceae with the highest ratio of repetitive sequences found so far. Phylogenetic analyses reveal that magnoliids are as sister to the eudicots-monocots clade, which provides insight into the phylogenetic position between eudicots and monocots. WGD analysis showed that an ancient duplication event occurred in the M. officinalis genome, is shared by Lauraceae. Based on gene functional annotations and RNA-seq, we identified several key enzyme-coding gene families associated with the biosynthesis of lignan in the genome. In addition, the floral fragrance of M. officinalis is mainly attributed to its terpenes, we classified many putative genes involved in the biosynthesis of terpene synthases. The construction of the M. officinalis genome sequence map will serve as a reference for further studies of Magnolia as well as other Magnoliaceae.

Venom gland is a highly efficient venom production system to maintain their predatory arsenal. Venom toxins mRNA has been shown to increase abruptly in snake after venom expenditure, while the dynamics of venom accumulation during synthesis are poorly understood. Here, PacBio long-read sequencing, Illumina RNA sequencing (RNA-seq), and label-free proteome quantification were used to investigate the composition landscape and time- and temperature-dependent dynamics changes of the Bungarus multicinctus venom gland system. Transcriptome data (19.5223 Gb) from six adult B. multicinctus tissues were sequenced using PacBio RS II to generate a reference assembly, and average 7.28 Gb of clean RNA-seq data was obtained from venom glands by Illumina sequencing. Differentially expressed genes (DEGs) mainly were protein processing rather than venom toxins. Post-translational modifications provided the evidence of the significantly different proportions of toxins in the venom proteome with the changing of replenishment time and temperature, but constant of venom toxins mRNA in the venom gland transcriptome. Dynamic of toxins and genes involved in venom gland contraction suggesting the formation of the mature venom gland system would take at least 9 days. In addition, 59 toxin processing genes were identified, peptidylprolyl isomerase B of which underwent positive selection in Toxicofera. These results provide a reference for determining the extraction time of venom, production of polyclonal and monoclonal antibody for precise treatment plans of venomous snakebites, and construction of an in vitro synthesis system for snake venom protein.

Aconitum scaposum Franch 1894 belongs to the Genus Aconitum and Subgenus Lycoctonum (Ranunculaceae). It is widely distributed in China and adjacent areas, used as herbal medicine and had highy toxic components. This species has little reasearch information, especially its chloroplast (cp) genome information being unclear. Therefore, with the method of high salt and low pH to extract the cp of A. scaposum, we sequenced and assembled the complete cp genome of A. scaposum using Illumina high-throughput sequencing platform. The results showed the cp genome of A. scaposum was 157 688 bp in length, including a pair of inverted repeated regions (IRa 26 156 bp and IRb 26 232 bp, respectively), large single copy region (LSC 69 309 bp) and small single copy region (SSC 16 917 bp). And cp genome of A. scaposum consisted of 145 unique genes, 8 ribosomal RNA (rRNA) genes and 38 transfer RNA (tRNA) genes, with GC content was 38%. Meanwhile, based on the cp complete genome, we performed the phylogenetic tree of 66 species with maximum likelihood (ML) method, respectively. Among them, we selected one Delphinium species as the outgroup and the bootstrap of each braches were greater than 90%. The results indicated that the phylogenetic relationship of A. scaposum was relatively closely related to A. scaposum var. vaginatum compared to other Aconitum species.

Magnoliae officinalis is the plant source of houpo, a widely used traditional Chinese medicine to treat symptoms of gastrointestinal diseases. Its main active components, magnolol (MG) and honokiol (HK), have excellent pharmacological actions, but little research has focused on the functional genes involved in the MG and HK metabolic pathways. In this study, using RNA-seq and gene expression profile, we present the first transcriptome characterization of M. officinalis leaves, twigs and stems. Based on similarity search against nonredundant protein databases, 30,660 contigs had at least a significant alignment to existing public database. Pathway analysis showed that 8707 contigs were assigned to 317 KEGG pathways. A second skeleton pathway with 14 putative homologous genes was also identified as involved in lignan biosynthesis. Expression profiles of these 14 genes showed that leaves and twigs seem to have higher transcript levels for lignan components than in stem tissue; this result was then verified by qRT-PCR. Our work will immensely facilitate metabolic research on lignan biosynthesis in M. officinalis.

Phoebe zhennan S. Lee et F. N. Wei (Lauraceae), is the main source of Gold Phoebe, a rare and extremely valuable wood in China. However it has undergone a dramatic decline. In this study, we used 12 amplified fragment length polymorphism primer combinations to assay 92 accessions, which were highly representative of the entire P. zhennan germplasm. It revealed that P. zhennan consisted of three genetic populations, named as SCZ (central Sichuan), CQH (eastern Sichuan, Chongqin, Hubei and Hunan) and YG (Yunnan and Guizhou), probably owing to natural selection caused by topography differences. The CQH population further diverged into two geographical sub-populations: CD-CQ (SCD and west region of Chongqin) and HB-HN (eastern side of Chongqin, Hubei and Hunan). The loci were moderately polymorphic (40.4%). The genetic distance between SCZ and YG was the highest, between CD-CQ and HB-HN the lowest. Pairwise fixation indices (Ф PT ) between any inferred populations were significant. This rare species exhibited low genetic diversity; therefore, the results provided significant data related to the conservation and management of P. zhennan . That is, with this genetic information, land managers are equipped with better tools allowing them to more effectively protect this species and its limited genetic diversity.

Isolation and Identification of Disease Pathogens in Plant Root

Coptis chinensis Franch. (Weilian in Chinese) is an important medicinal plant used in traditional Chinese medicines. The identification of habitats associated with good quality plant material is a challenge. In this study, we determined 59 samples from 12 different habits. Other than the samples from Zhenping, the content of six selected alkaloids in C. chinensis did not differ significantly among the habits. Furthermore, the results of the genetic analysis showed that the genetic diversity and the genetic distance among the samples were low, suggesting that the C. chinensis samples from different habits had the same genetic characteristics. These results would suggest that the quality of the drugs are not influenced by the habitats the plant is growing in.

Breast cancer is one of the most frightful causes of death among females worldwide. Accumulating evidence attached the importance of microRNAs negative regulation to tumorigenesis in breast cancer, suggesting novel cancer therapies targeting microRNAs modulation. Recent studies demonstrated that isoliquiritigenin could inhibit breast cancer cells proliferation and migration, but the underlying mechanism is still limited. In this study, the anti-cancer effects as well as the detailed mechanisms of isoliquiritigenin were explored. The results proved that isoliquiritigenin could negatively regulate breast cancer growth through the induction of apoptosis. We also verified the anti-cancer effect of isoliquiritigenin on migration and invasion, and identified highly expressed miR-374a as one of the main microRNAs down-regulated by isoliquiritigenin treatment in breast cancer. Further study displayed that isoliquiritigenin increased PTEN expression through the decrease of miR-374a expression to inhibit the aberrant Akt signaling. Our findings suggest isoliquiritigenin as a novel anti-cancer candidate significantly regulating miR-374a/PTEN/Akt axis in microRNA-based breast cancer therapies.

High chilling-susceptibility is becoming the bottleneck for cultivation and commercialization of Jatropha curcas L. For insights to chilling resistance ability of this plant species, a cold response transcription factor, JcCBF2, was cloned and studied. It codes a 26 kDa protein, which contains all conserved motifs unique to the C-repeat binding factor (CBF) family and has high similarity to CBFs of Ricinus communis and Populus. Its transcripts express specifically in leaves of Jatropha at cold temperature. After transmitting the report vector, 35S::JcCBF2-GFP, into Arabidopsis thaliana, JcCBF2 protein is main detected in cell nucleus, being consistent to the nuclear orientation signal in its N-terminal. Compared to the control Arabidopsis, the frozen leaves of JcCBF2-overexpressed seedlings grow stronger with less malondialdehyde, smaller leaf conductivity and activer superoxide dismutase, showing their higher freezing tolerance. RT-PCR tests revealed that JcCBF2 functioned mainly at the early stage (0-6 h) of resistance events in Arabidopsis, and its transcripts reduced after 6 h. In addition, JcCBF2 could quickly regulate transcripts of some cold-responsive (COR) genes such as RD29A, COR105A and COR6.6, also during the early stage of frozen treatment. This study not only proves the chilling resistance roles of JcCBF2, but also presents a candidate gene engineering for improvement of chilling tolerance in J. curcas.

A putative fatty acyl-acyl carrier protein (acyl-ACP) thioesterase (thioesterase) full-length cDNA sequence named as ClFATB1 was obtained from the seed cDNA library of Cinnamomum longepaniculatum by the SMART-RACE method. The novel gene encodes a protein of 382 amino acid residues with close homology to fatty acid thioesterase type B (FATB) enzymes of other plants, with two essential residues (His285 and Cys320) for thioesterase catalytic activity. The gene was transcribed in all tissues of C. longepaniculatum, the highest being in seeds. Recombinant ClFATB1 in Escherichia coli had higher specific activities against saturated 16:0- and 18:0-ACPs than on unsaturated 18:1-ACP. Overexpression of ClFATB1 in transgenic tobaccos upregulated thioesterase activities of crude proteins against 16:0-ACP and 18:0-ACP by 20.3 and 5.7%, respectively, and resulted in an increase in the contents of palmitic and stearic acids by 15.4 and 10.5%, respectively. However, ectopic expression of this gene decreased the substrate specificities of crude proteins to unsaturated 18:1-ACP by 12.7% in transgenic tobacco and lowered the contents of oleic, linoleic, and linolenic acids in transgenic leaves. So ClFATB1 would potentially upregulate the synthesis of saturated fatty acids and downregulate unsaturated ones in the fatty acid synthesis pathway of plants.

Desire is a key concept in Deleuzian philosophy. Deleuze's desire is quite different from that of other thinkers. Both in the West and in China, in the past as well in the present, desire is usually understood as something abnormal, avaricious and excessive, the opposite of rationality, to be controlled and suppressed in man. Deleuze's desire is much wider, referring not only to man, but also to animals, objects and social institutions. In Deleuze's view, desire is not a psychic existence, not lack, but an active and positive reality, an affirmative vital force. Desire has neither object, nor fixed subject. It is like labour in essence, productive and actualisable only through practice. This paper discusses the formation, evolution and limitation of Deleuzian desire.

An MSAP analysis method was established for detecting DNA methylation of Aconitum carmichaeli leaves, and the DNA methylation of different leaf shapes and different leaf position was analyzed by MSAP. The study made experiments on the leaves of different position of mosaic and moxa leaf type A. carmichaeli, researched the effects of restriction digestion of genomic DNA by using two restriction enzymes, screened the suitable selective amplification primers, and analyzed the methylation differences of leaves by calculating the 6% acrylamide gel electrophoresis bands and lane. The best reaction system of MSAP was obtained, under the conditions of 37 ℃, the 16 h incubated time was more suitable for 150 ng DNA, and 25 pairs of selective amplification primers were selected from 256 pairs. Totally, 273 electrophoresis bands were obtained by 25 pairs of selective primers, including 228 non methylation or single chain methylation bands,27 double chain methylation bands,and 18 single stranded methylation bands, the total methylation rate was 16.48%. The methylation rate was slightly different in mosaic and moxa leaf type A. carmichaeli leaf, which were 15.36%, 14.34%, respectively, and article 8, article 6 nucleotide fragments of genome methylation modification differences were obtained, accounted for 3%, 2.26% of the total number of bands. Based on this study it can provide new ideas for molecular identification, breeding and cultivation, and genetic evolution of A. carmichaeli.

Herein, we cloned a full-length cDNA encoding allene oxide cyclase (AOC, EC 5.3.99.6) that is a key enzyme in jasmonates (JAs) biosynthetic pathway from Jatropha curcas L., an important plant species as its seed is the raw material for biodiesels, named as JcAOC (GenBank accession no. FJ874630). The cDNA was 924 bp in length with a complete open reading frame of 750 bp, which encoded a polypeptide of 250 amino acids including a putative signal peptide of 65 amino acid residues and a mature protein of 185 amino acids with a predicted molecular mass of 20.7 kDa and a isoelectric point of 6.24. Phylogenetic analysis indicated that JcAOC belonged to the AOC superfamily. Semi-quantitative RT-PCR analysis revealed that JcAOC mRNA was expressed in roots, stems, leaves, young seeds, endosperms, and flowers, but that the expression level was highest in leaves and lowest in seeds, and mRNA expression of JcAOC could be induced by salt stress (300 mM NaCl) and low temperature (4°C). Furthermore, the full-length coding region of JcAOC excluding signal peptide sequence was inserted into pET-30a and was successfully expressed in Escherichia coli. Overexpression of JcAOC in E. coli conferred its resistance to salt stress and low temperature.

Phospholipase D (PLD, EC 3.1.4.4) is a key enzyme involved in phospholipid catabolism, initiating a lipolytic cascade in membrane deterioration during senescence and stress, which was cloned from Jatropha curcas L., an important plant species as its seed is the raw material for biodiesels. The cDNA was 2,886 bp in length with a complete open reading frame of 2,427 bp which encoded a polypeptide of 808 amino acids including a putative signal peptide of 53 amino acid residues and a mature protein of 755 amino acids with a predicted molecular mass of 86 kD and a pI of 5.44, having two highly conserved HKD' motifs. Phylogenetic analysis indicated the J. curcas PLD alpha (JcPLDalpha) showed a high similarity to other PLD alpha from plants. Semi-quantitative RT-PCR analysis revealed that it was especially abundant in root, stem, leaf, endosperm and flower, weakly in seed. And the JcPLDalpha was increasedly expressed in leaf undergoing environmental stress such as salt (300 mM NaCl), drought (30% PEG), cold (4degreeC) and heat (50degreeC). The JcPLDalpha protein was successfully expressed in Escherichia coli and showed high enzymatic activities. Maximal activity was at pH 8 and 60degreeC.

Ribosome-inactivating proteins (RIPs) represent a type of protein that universally inactivates the ribosome thus inhibiting protein biosynthesis. Curcin-L was a type I RIP found in Jatropha curcas L.. Its expression could be activated in leaves by treatments with abscisic acid, salicylic acid, polyethylene glycol, temperature 4, 45 degrees C and ultraviolet light. A 654 bp fragment of a 5' flanking region preceding the curcin-L gene, designated CP2, was cloned from the J. curcas genome and its expression pattern was studied via the expression of the beta-glucuronidase (GUS) gene in transgenic tobacco. Analysis of GUS activities showed that the CP2 was leaf specific, and was able to drive the expression of the reporter gene under stress-induction conditions. Analysis of a series of 5'-deletions of the CP2 suggested that several promoter motifs were necessary to respond to environmental stresses.

The complete chloroplast genome sequence of an important medicinal plant of the family Ranunculaceae, Aconitum carmichaelii Debx., was characterized in this study. The assembled chloroplast genome was 154,776 bp in length, which included a large single-copy (LSC), a small single-copy (SSC), and two inverted repeat (IR) regions of 86,330bp, 15,986 bp, and 26,193 bp, respectively. The GC content of the genome was 38.1%. Phylogenetic analysis with the whole nucleotide sequences of reported Aconitum chloroplast genomes indicated a close relationship of A. carmichaelii with A. kusnezoffii.

A PAL gene designated as JcPAL1 was cloned from J. curcas L. The full-length is 2336 bp in size with one intron and two exons, encoding a polypeptide of 713 amino acids. Its 5'-upstream region is rich in putative cis-elements including not only PAL typical TATA box, L-box and transcriptional initiation site (TIS) but also light responding motifs. Expression pattern analysis indicated that JcPAL1 were expressed in all tissues, most highly in flowers. When Treated with ABA, GA3, high and low temperature, expression of JcPAL1 were induced. Recombinant JcPAL1 has a pH optimum at 8.7 and a temperature optimum at 60°C in 100 mM Tris-HCl buffer. The Km and Kcat values are 0.125 mM and 1.73 S(-1) for L: -phenylalanine, and 1.312 mM and 0.109 S(-1) for L: -tyrosine, respectively. These findings suggested that JcPAL1 might involve in the J. curcas responding to various stresses and L: -Phe should be its true physiological substrate. This study is essential prior to uncover whether and how the PAL initiated phenylpropanoid metabolic networks functioning in the defense responses of J. curcas.

To date, two types of ribosome-inactivating proteins (RIPs) have been found in Jatropha curcas. One is curcin, which has been isolated from the endosperm, and the other is curcin-L, which is expressed in leaves upon stress treatment. Phylogenetic analysis of the predicted amino acid sequences of the RIPs in plants revealed that these belong to a major subfamily and are close to trichosanthin (TCS). Studies on the mRNA and protein levels showed that both curcin and curcin-L have an organ-specific expression pattern. Curcin is only expressed and accumulated in the endosperm; its expression begins in the globular embryo period and peaks during the mature embryo period. In contrast, curcin-L is only expressed in the leaves, but its expression is induced by certain conditions such as treatment with phytohormones or polyethylene glycol, exposure to high and low temperatures, and fungal infection. Analysis of the 5’ flanking regions of curcin and curcin-L revealed that the 5’ flanking region of curcin-L has three major inserted fragments, which are not present in the corresponding region of curcin. Comparison of characteristic cis-elements suggests the presence of several motifs that are involved in the endosperm-specific expression in the 5’ flanking region of curcin, while in curcin-L some stress- and defense-responsive motifs are found to be mainly located in the three inserted fragments. Comparison of the antifungal activity of the two RIPs showed that the one of curcin-L is higher than that of curcin. Differences in the expression and activity of curcin and curcin-L suggest that these two RIPs have different functions.

A cDNA encoding translationally controlled tumor protein (TCTP) of Jatropha curcas L., JcTCTP, was isolated from an endosperm cDNA library. JcTCTP consisted of a 5' untranslated region (UTR) of 526 bp, a 3' UTR of 377 bp and an open reading frame (ORF) of 507 bp, encoding a protein of 168 amino acid residues, which contained two signature sequences of TCTP family. Its deduced amino acid sequence was similar to the other known plants TCTPs in a range of 77.4-92.3%. Expression of JcTCTP was the highest in the stem, endosperm at embryo formation stage and embryo of J. curcas tissues, and the lowest in the endosperm at seminal leaf embryo stage and flower, demonstrating a pattern of temporal and spatial specific expression.