Terpenoids are important secondary metabolites of plants that possess both pharmacological activity and economic value. Terpene synthases(TPSs) are key enzymes in the synthesis process of terpenoids. In order to investigate the TPS gene family members and their potential functions in Schizonepeta tenuifolia, this study conducted a systematic analysis of the TPS gene family of S. tenuifolia based on the whole genome data of S. tenuifolia using bioinformatics methods. The results revealed 57 StTPS members identified from the genome database of S. tenuifolia. The StTPS family members encoded 285-819 amino acids, with protein molecular weights ranging from 32.75 to 94.11 kDa, all of which were hydrophilic proteins. The StTPS family members were mainly distributed in the cytoplasm and chloroplasts, exhibiting a random and uneven physical localization pattern. Phylogenetic analysis showed that the StTPS genes family were divided into six subgroups, mainly belonging to the TPS-a and TPS-b subfamilies. Promoter analysis predicted that the TPS gene family members could respond to various stressors such as light, abscisic acid, and methyl jasmonate(MeJA). Transcriptome data analysis revealed that most of the TPS genes were expressed in the roots of S. tenuifolia, and qRT-PCR analysis was conducted on genes with high expression in leaves and low expression in roots. Through the analysis of the TPS gene family of S. tenuifolia, this study identified StTPS5, StTPS18, StTPS32, and StTPS45 as potential genes involved in sesquiterpene synthesis of S. tenuifolia. StTPS45 was cloned for the construction of an prokaryotic expression vector, providing a reference for further investigation of the function and role of the TPS gene family in sesquiterpene synthesis.
Phylogeny ; Terpenes/metabolism* ; Plant Proteins/metabolism* ; Lamiaceae/genetics* ; Sesquiterpenes

OBJECTIVEWith the purpose of selecting adequate quality and high production of Schizonepeta tenuifolia, the comparative experiments were carried out on different strain of S. tenuifolia in 2007.

METHODThe test fields were divided into blocks randomly, and the agronomy characters were investigated in harvest time; the content of volatile oil was measured by steam distillation and the pulegone were determined by HPLC.

RESULTThe yield of S4 was 18.63% and 29.99% higher than that of CK1 and CK2, respectively. The contents of volatile oil and pulegone were also higher than those of CK and other strains in this test.

CONCLUSIONS4 shows the advantages of high production, strong disease resistance and high active components. S4 would be extended as the good breed in production.

Agriculture ; Breeding ; Lamiaceae ; chemistry ; genetics ; growth & development ; physiology ; Quality Control ; Time Factors ; Volatilization

Abstract: In order to enhance the content of secondary metabolites patchouli alcohol in Pogostemon cablin, we induced polyploid hairy roots and their plant regeneration, and determined the content of patchouli alcohol through artificial chromosome doubling with colchicine. The highest rate of polyploidy induction was more than 40% when hairy roots were treated with 0.05% colchicine for 36 h. The obtained polyploid hairy roots formed adventitious shoots when cultured in an MS medium with 6-BA 0.2 mg/L and NAA 0.1 mg/L for 60 d. Compared with the control diploid plants, the polyploid hairy root-regenerated plants of P. cablin had more developed root systems, thicker stems, shorter internodes and longer, wider and thicker leaves. Observation of the chromosome number in their root tip cells reveals that the obtained polyploid regenerated plants were tetraploidy, with 128 (4n = 128) chromosomes. The leaves contained around twice as many stomatal guard cells and chloroplasts as the controls, but the stomatal density declined with increasing ploidy. The stomatal density in diploid plants was around 1.67 times of that in polyploid plants. GC-MS analysis shows that the content of patchouli alcholol in the hairy root-derived polyploid plants was about 4.25 mg/g dry weight, which was 2.3 times of that in diploid plants. The present study demonstrates that polyploidization of hairy roots can stimulate the content of patchouli alcholol in medicinal plant of P. cablin.
Colchicine ; Diploidy ; Lamiaceae ; genetics ; growth & development ; Plant Roots ; growth & development ; Plants, Medicinal ; genetics ; growth & development ; Polyploidy ; Regeneration ; Sesquiterpenes ; chemistry

Hd-Zip, a unique transcription factor in plant kingdom, influences the growth, development, and secondary metabolism of plants. Hd-zip Ⅳ is thought to play an important role in trichome development of Schizonepeta tenuifolia. This study aims to explore the functions of StHD1 and StHD8 in Hd-zip Ⅳ subfamily in peltate glandular trichome development. To be specific, the expression patterns of the two genes and interaction between the proteins encoded by them were analyzed based on transcriptome sequencing and two-hybrid screening. The subcellular localization was performed and functions of the genes were verified in tobacco and S. tenuifolia. The results showed that StHD1 and StHD8 had high similarity to HD-Zip Ⅳ proteins of other plants and they all had the characteristic conserved domains of HD-Zip Ⅳ subfamily. They were located in the nucleus. The two genes mainly expressed in young tissues and spikes, and StHD1 and StHD8 proteins interacted with each other. The density and length of glandular trichomes increased significantly in tobacco plants with the overexpression of StHD1 and StHD8. Inhibiting the expression of StHD1 and StHD8 by VIGS(virus-induced gene silencing) in S. tenuifolia resulted in the reduction in the density of peltate glandular trichomes, the expression of key genes related to mono-terpene synthesis, and the relative content of limonene and pulegone, the main components of monoterpene. These results suggested that StHD1 and StHD8 of S. tenuifolia formed a complex to regulate glandular trichomes and affect the biosynthesis of monoterpenes.
Trichomes/metabolism* ; Lamiaceae/genetics* ; Tobacco/genetics* ; Monoterpenes/metabolism* ; Cloning, Molecular ; Plant Proteins/metabolism* ; Gene Expression Regulation, Plant

OBJECTIVETo study the genetic polymorphism and intraspecific genetic differentiation of different populations of Pogostemon cablin, and find out the effective method to distinguish DNA fingerprint of different populations of P. cablin.

METHODFive plant populations of P. cablin were analyzed by RAPD markers. PopGen 32 software for clustering analysis and calculating. Fourteen of the 80 random primers were tested to possess the stronger detecting effect of polymorphous character.

RESULTA total of 84 bands was amplified by the 10 primers, among them 17 bands were monomorphic. 67 of them were polymorphic. The results indicated that the genetic variations existed within the different plant populations of the same species.

CONCLUSIONIt is feasible by RAPD technique with specifically primer to analyze the genetic diversity and identify 5 plant populations of P. cablin. RAPD technique has provided a new path for identification and classification of P. cablin genetic germplasm.

China ; Cluster Analysis ; DNA Fingerprinting ; DNA, Plant ; genetics ; Ecosystem ; Lamiaceae ; genetics ; Phylogeny ; Plants, Medicinal ; genetics ; Polymorphism, Genetic ; Random Amplified Polymorphic DNA Technique

AIMTo analyze sequences of the nuclear ribosomal RNA small subunit (18S rRNA) gene and the chloroplast matK gene of crude drug Patchouli (Pogostemon cablin) in order to provide molecular evidence for identification of Patchouli drug.

METHODSTo sequence the entire 18S rRNA gene and partial matK gene of Patchouli from Guangzhou and its substitute Wrinkled Gianthyssop (Agastache rugosa) from Sichuan using PCR direct sequencing and to detect the homology of two gene sequences between these two crude drugs.

RESULTSThe complete 18S rRNA gene sequence is 1,805 bp in length for Patchouli from Guangzhou whereas 1,794 bp for Wrinkled Gianthyssop from Sichuan. The 3'-end sequence of matK gene is 521 bp (747-1,268 nt from upstream of matK gene) for these two crude drugs. Based on multiple sequence alignment, it is found that there are 18 variable sites and 11 aligned gap sites in 18S rRNA sequence, 49 variable sites in 3'-matK sequence between these two crude drugs. The homology is 98.4% for 18S rRNA and 90.6% for 3'-matK between two crude drugs, respectively.

CONCLUSIONDNA sequencing can provide an accurate and reliable tool in the crude drug identification of Patchouli and its substitute Wrinkled Gianthyssop.

Agastache ; genetics ; Base Sequence ; DNA, Plant ; analysis ; Drug Contamination ; Lamiaceae ; classification ; genetics ; Molecular Sequence Data ; Plant Stems ; genetics ; Plants, Medicinal ; genetics ; RNA, Ribosomal, 18S ; analysis ; genetics ; Sequence Alignment ; Sequence Analysis, DNA ; Sequence Homology ; Species Specificity

AIMTo provide molecular evidence for quality evaluation and GAP production of Pogostemon cablin (Blanco) Benth. cultivated in different regions in Guangdong and Hainan provinces, China, by comparing two sequences (1.2 kb of plastid matK gene and 1.8 kb of nuclear 18S rRNA gene) and two chemotypes (Pogostone-type and Patchouliol-type in essential oil composition).

METHODSPCR direct sequencing was applied to detemine the matK and 18S rRNA sequences for six samples of Pogostemon cablin from different localities.

RESULTSThe matK sequences of six samples of Pogostemon cablin from different regions of cultivation are 1,245 bp in length, which coding 415 amino acids of protein (maturase), and 18S rRNA sequences are 1,803-1,805 bp in size. Based on multiple sequence alignment, there are 47 variable sites in the matK sequence of these six samples, 17 in the 18S rRNA sequence. The cluster tree reconstructed by UPGMA method shows that the sequence divergence both in matK and 18S rRNA genes among six samples of Pogostemon cablin was well correlative with their regions of cultivation and intraspecific chemotypes of essential oil composition.

CONCLUSIONCombining with chemical and biogeographical data, DNA sequencing can become a powerful tool in the key technique-species identification of quality evaluation and GAP production of Pogostemon cablin.

Base Sequence ; DNA, Plant ; analysis ; Endoribonucleases ; genetics ; Genes, Plant ; Lamiaceae ; chemistry ; classification ; genetics ; Molecular Sequence Data ; Nucleotidyltransferases ; genetics ; Oils, Volatile ; analysis ; Polymorphism, Genetic ; Quality Control ; Sequence Analysis, DNA ; Species Specificity

To investigate the correlation between genotype and distribution of Pogostemon cablin by sequencing ITS1 and ITS2 genes, and provide molecular information for its germplasm evaluation, ITS1 and ITS2 genes of Pogostemon cablin from different localities were identified by PCR direct sequencing. The sequences of ITS1 and ITS2 genes were 424 bp and 380 bp in length, respectively. And nineteen base substitutions were observed in ITS1 gene, and five in ITS2 gene. The results showed a good correlation between genotype and distribution of Pogostemon cablin, and ITS gene sequencing could provide useful molecular information for germplasm evaluation of the plant species verification.
Base Sequence ; Biodiversity ; China ; Cluster Analysis ; DNA, Plant ; chemistry ; genetics ; DNA, Ribosomal ; chemistry ; genetics ; DNA, Ribosomal Spacer ; chemistry ; genetics ; Genotype ; Geography ; Lamiaceae ; classification ; genetics ; growth & development ; Molecular Sequence Data ; Phylogeny ; Plant Leaves ; genetics ; Plants, Medicinal ; classification ; genetics ; growth & development ; Sequence Analysis, DNA

OBJECTIVETo observe the effect of volatile oil of Schizonepetae Herba (VOSH), and its essential components-menthone and pulegone against anti-influenza virus A/PR/8/34 (H1N1) in vivo and in vitro, as well as the signaling mechanism of its toll-like receptor/interferon (TLR/IFN).

METHODThe lung-adapted PR-8 virus model was prepared in mice. They were administered with preventive and therapeutic drugs, and the hemagglutination titer of model animals was determined to evaluate in vivo effect against H1N1. ELISA test was conducted to observe the effect on IFN-alpha, IFN-beta, IL-2, IL-6 and TNF-alpha in serum, as well as IFN-beta secretion in H1N1 infected MDCK supernatant. Real-time RT-PCR was employed to observe the expression levels of IRAK4 and TLR3 mRNA.

RESULTThe in vivo experiment shows that the hemagglutination titer was significantly decreased when the mice were treated with VOSH (0.266 mg x kg(-1)), menthone(0.5 mg x kg(-1)) and pulegone (0.19 mg x kg(-1)) in therapeutic way; VOSH (0.226 mg x kg(-1)) had a significant effect on increasing serum levels of IFN-alpha, IL-2; Methone (0.5 mg x kg(-1)) had a significant effect on increasing serum levels of IFN-beta; Methone (0.5 mg x kg(-1)) and pulegone (0.19 mg x kg(-1)) had a significant effect on decreasing serum levels of IL-6; VOSH (0.452, 0.226 mg x kg(-1)) and pulegone (0.19 mg x kg(-1)) had a significant effect on decreasing serum levels TNF-alpha. The in vitro experiment showed that the expression levels of IRAK4 mRNA and IFN-beta were significantly increased in VOHS (0.1 g x L(-1)) and pulegone (0.1 g x L(-1)) groups; and the menthone (0.25 g x L(-1)) group showed a significant rise in the expression levels of IRAK4 mRNA, but a notable decline in TLR3 mRNA.

CONCLUSIONThe administration with VOSH, methone and pulegone in therapeutic way can significantly decrease the hemagglutination titer, which demonstrates the anti-virus effect of the administration in therapeutic way, but no notable efficacy of the administration in preventive way. The in vivo anti-virus mechanism is related to regulation of IFN-alpha, IFN-beta and IL-2.

Animals ; Drugs, Chinese Herbal ; pharmacology ; Female ; Humans ; Influenza A Virus, H1N1 Subtype ; drug effects ; physiology ; Influenza, Human ; drug therapy ; genetics ; immunology ; virology ; Interferon-alpha ; genetics ; immunology ; Interleukin-1 Receptor-Associated Kinases ; Interleukin-2 ; genetics ; immunology ; Interleukin-6 ; genetics ; immunology ; Lamiaceae ; chemistry ; Male ; Mice ; Oils, Volatile ; pharmacology ; Tumor Necrosis Factor-alpha ; genetics ; immunology

OBJECTIVETo study the effects of volatie oil of Schizonepeta tenuifolia Briq herb and Saposhnikovia divaricata Schischke root (OSS) on proinflammatory cytokine expression and regulation in rats.

METHODOA and LPS were injected intravenously to rats to develop acute lung injury (ALI). The rats were treated with OSS (45.19 microL kg(-1)). The pathological sections of lung tissue were prepared and observed in acute lung injury rats. The expression of nuclear factor-kappa B p65 (NF-kappaB p65), intercellar adhesion molecule CD54, and NF-kappaB p65 mRNA were determined in lung cells.

RESULTvolatie oil of Schizonepeta tenuifolia Briq herb and Saposhnikovia divaricata Schischke root significantly inhibited the expression of CD54, the activation of NF-kappaB p65, and the transcription of NF-kappaB p65 mRNA.

CONCLUSIONOSS can reduce the expression of CD54 and NF-kappaB p65 protein synthesis, which may be its anti-inflammatory molecular mechanisms.

Animals ; Anti-Inflammatory Agents ; isolation & purification ; pharmacology ; Apiaceae ; chemistry ; Drug Combinations ; Gene Expression Regulation ; drug effects ; Immunohistochemistry ; Intercellular Adhesion Molecule-1 ; biosynthesis ; Lamiaceae ; chemistry ; Lipopolysaccharides ; Male ; Oils, Volatile ; isolation & purification ; pharmacology ; Oleic Acid ; Plant Oils ; isolation & purification ; pharmacology ; Plants, Medicinal ; chemistry ; RNA, Messenger ; biosynthesis ; genetics ; Random Allocation ; Rats ; Rats, Wistar ; Respiratory Distress Syndrome, Adult ; chemically induced ; metabolism ; prevention & control ; Transcription Factor RelA ; biosynthesis ; genetics