Objective To study the chemical constituents in the leaves of Lindera aggregate.MethodsVarious column chromatographic techniques were used to separate and purify the chemical constituents and their structures were elucidated by spectral analyses.Results Nine compounds were isolated and identified as quercetin(Ⅰ),and kaempferol(Ⅱ),avicularin(Ⅲ),afzelin(Ⅳ),dihydrokaempferol(Ⅴ),astragaline(Ⅵ),kaempferol-3-O-?-D-xylopyranoside(Ⅶ),juglalin(Ⅷ),and kaempferol-3-O-(2″-O-?-D-glucopyranosyl)-?-L-rhamnopyranoside(Ⅸ).Conclusion Compounds Ⅲ—ⅠⅩ are isolated from the title plant for the first time.Compounds Ⅴ—Ⅸ are isolated from the plants of Lindera Thunb.for the first time.

Cytochrome P450 (CYP450) is a key element in the Ganoderma triterpenoid biosynthetic pathway. The catalytic reaction process for CYP450 requires NADPH / NADH for electron transfer. After searching the genome dataset of Ganoderma lucidum, the unique sequence encoding CYP450 and NADPH were discovered, separately. The open reading frames of GLCYP450 and GLNADPH were cloned separately using RT-PCR strategy from G lucidum. The appropriate restriction enzyme cutting sites were introduced at the 5' and 3' ends of gene sequence. The genes of GLCYP450 and GLNADPH were recombined into the yeast expression vector pESC-URA, leading to the formation of the yeast expression plasmid pESC-GLNADPH-GLCYP450. This study provides a foundation for researching Ganoderma triterpene biosynthesis using the approach of synthetic biology.

3-Hydroxy-3-methylglutaryl coenzyme-A reductase (HMGR), the first enzyme of mavalonic acid pathway, is one of the key devices involved in ginsenoside biosynthesis based on synthetic biology approach. The open reading frame of a novel HMGR gene from Panax ginseng (PgHMGR2) was cloned and analyzed in this study. PgHMGR2-encoding protein showed 71.6% sequence similarity to a P. ginseng HMGR in GenBank. The full-length cDNA sequence of PgHMGR2 containing 1 770 bp, which encodes 589 amino acids, was cloned by RT-PCR strategy from P. ginseng. The bioinformatic analysis showed that PgHMGR2-encoding protein contained two transmembrane regions and the HMG_CoA_reductase domain, without signal peptide. The protein sequence of PgHMGR2 had the highest sequence similarity (99%) with Panax quinquefolius HMGR (GenBank accession No. ACV65036). The expression level of PgHMGR2 was the highest in flower based on a real-time PCR analysis, followed by leaf and root, and the lowest was in stem. The result will provide a foundation for exploring the molecular function of PgHMGR2 involved in ginsenoside biosynthesis based on synthetic biology approach in P. ginseng plants.

DNA barcoding is a technique in which species identification and discovery are performed by using short and standard fragments of DNA sequences. In this study, eleven species of Paris, including seven varieties, were sampled. Five chloroplast sequences, psbA-trnH, rpoB, rpoC1, rbcL, matK, and one nuclear marker, the second internal transcribed spacer (ITS2) of ribosomal DNA, were amplified and sequenced. The PCR amplification and sequencing efficiency, intra- and inter-specific divergence and barcoding gap were used to evaluate different loci, and the identification efficiency was assessed using BLAST1 and Nearest Distance methods. The ITS2 sequences in the studied samples of Paris were amplified and sequenced successfully using primers designed by our group, while matK showed low level in the amplification and psbA-trnH was difficult for sequencing because of over 800 bp and poly (A) structure. Analysis of the intra- and inter-specific divergence and barcoding gap showed ITS2 was superior to other loci. The ITS2 showed a much higher percentage of success (100%) in identification than other five loci, none of which indicated more than 50% except matK (52.9%). The 2-locus combination of rbcL+matK didn't improve ability of authentication. In addition, the rate of successful identification with ITS2 kept 100% when the samples were expanded to 67 samples of 29 species. In conclusion, ITS2 can be used to correctly identify medicinal plants of Paris, and it will be a potential DNA barcode for identifying medicinal plants of other taxa.

To investigate the profile of gene expression in Salvia miltiorrhiza and elucidate its functional gene, 454 GS FLX platform and Titanium regent were used to produce a substantial expressed sequence tags (ESTs) dataset from the root of S. miltiorrhiza. A total of 46 722 ESTs with an average read length of 414 bp were generated. 454 ESTs were combined with the S. miltiorrhiza ESTs from GenBank. These ESTs were assembled into 18 235 unigenes. Of these unigenes, 454 sequencing identified 13 980 novel unigenes. 73% of these unigenes (13 308) were annotated using BLAST searches (E-value < or = 1e-5) against the SwissProt, KEGG TAIR, Nr and Nt databases. Twenty-seven unigenes (encoding 15 enzymes) were found to be involved in tanshinones biosynthesis, and 29 unigenes (encoding 11 enzymes) involved in phenolic acids biosynthesis. Seventy putative genes were found to encode cytochromes P450 and 577 putative transcription factor genes. Data presented in this study will constitute an important resource for the scientific community that is interested in the molecular genetics and functional genomics of S. miltiorrhiza.

Chloroplast genome sequences have comprehensive application prospects in DNA barcoding and chloroplast engineering in traditional Chinese medicine. The complete chloroplast genome of Magnolia officinalis sequenced by high-throughput pyrosequencing and a sequencing procedure was established. Fourteen contigs were obtained after de nove assembly. The sequencing percent of coverage was 99.99%. The chloroplast genome is 160 183 bp in size, and has a typical quadripartite structure with the large (LSC, 88 210 bp) and small copy (SSC, 18 843 bp) regions separated by two copies of an inverted repeat (IRs, 26 565 bp each). chloroplast genes were successfully annotated, of which 17 genes located in each IR region. The chloroplast genome features in Magnolia officinalis are nearly identical to those from other Magnoliid chloroplast genomes. Phylogenetic analyses were performed based on 81 shared coding-genes for a total of 9 Magnolia samples of 5 closely related species. Results showed that distinguishing among species was generally straightforward at the species and population level. This study confirmed the effectiveness of our chloroplast genome sequencing procedure. The chloroplast genome can provide distinguishing differences to help identify Magnolia officinalis and its closely related plants.

ERF family transcription factor (TF) represented ethylene-responsive protein which harbored a conserved AP2 domain. After searching the plant transcription factor database, a total of 75 unigenes was found which contained AP2 domain from the transcriptome dataset of Panax quinquefolius L. One unique sequence of ERF transcript, named as PqERF1, was cloned with entire open reading frame of 933 base pairs (bp). Protein prediction result indicated that the gene was localized in nucleus and had a conserved AP2 domain. PqERF1 gene could be induced by methyl jasmonate (MeJA) which was consistent to the inducing profile of triterpene ginsenosides. InterproScan prediction indicated that PqERF1 was probably a pathogenesis-related gene. Sequence alignment and phylogenetic analysis demonstrated PqERF1 was with high identity and had relative close relationship to the NtERF4 (Nicotiana tabacum), PhERF12 (Petunia x hybrida) and DcERF1 (Daucus carota) which was related to plant defense, regulation of secondary metabolism and the flower senescence respectively. Therefore, the gene was likely involved in regulation of secondary metabolism, plant defense and physical processes which would provide gene resource for further study on secondary metabolite synthesis and molecular breeding of P. quinquefolius.

To identify the original plant of Daturae Flos from its adulterants by DNA barcoding, the sequences of ITS2, psbA-trnH, matK, rbcL of four species including Datura metel, Darura innoxia, Darura stramonium and Brugmansia arborea were compared and analyzed. The PCR and sequencing success rate of the four regions (ITS2, psbA-trnH, matK, rbcL) was 100%, 90%, 100% and 85%, respectively. Sequences were assembled with CodonCode Aligner. K2P distances were calculated and NJ tree was performed by MEGA 4.1. Thirty SNPs were found among ITS2 sequences, and 33 insert/deletes were found among psbA-trnH intergenic regions. The interspecific K2P distance of ITS2 and psbA-trnH was obviously higher than that of the intraspecific one. As to matK and rbcL, there was no "Barcoding Gap" existing between inter- and intra-specific distances. The NJ trees of the four regions/combinations were built separately. Samples of Brugmansia arborea were clustered into one clade, and the other species of Datura L. formed another clade. The results showed that either ITS2 or psbA-trnH was useful to identify Daturae Flos from its adulterants.

Objective Based on the principles of laser radiation protection, medical metroiogy and photoelectron technology, an automatic verification device and testing technology to provide verification and performance evaluation for laser protective spectacles and equipments which have the various functions in laser protection have been developed and established. Methods The current system comprises laser source, laser measuring instruments, software of computer detection and control and modules of optics and mechanics used in auxiliary equipment. By use of the verification device and the special test-recording method to be designed and studied by us, the measured data can be automatically acquired, processed, recorded, saved and printed and, furthermore, many parameters on protective performance of the measured laser protective spectacles can be tested and given. Results Laser wavelength of the verification apparatus are 1 064 nm and 532 nm, response range of wavelength is from 0.4 m to 1.1 μm, measuring range of laser energy is from 10-8 J to 0.3 J, measuring range of optical density for laser protective spectacles is from 0.1 to 8.0, stability is 0.21%, measuring uncertainty is 5%(k=2). Conclusion The automatic verification device is steady and reliable. The achieved performance indexes accord with the requirement of national standard on laser protection.

Objective To explore the clinical application value of early bundle therapy in patients with septic shock after per‐cutaneous nephrolithotomy(PCNL) .Methods The retrospective analysis was conducted patients with septic shock after PCNL ad‐mitted to the central ICU of the First Affiliated Hospital ,University of South China from January 1st ,2011 to september 30 ,2013 . The patients were divided into non‐bundle therapy group and bundle therapy group according to whether treated by early bundle therapy .the APACHE‐Ⅱscore and SOFA score in the before and 1 ,3 ,7 d after treatment ,mortality rate within 28 d and length of ICU were compared with both groups .Results 54 patients were enrolled in the study ,there were 28 and 26 patients in non‐bundle therapy group and bundle therapy group ,respectively .The clinical data of patients in both groups had no significant difference be‐tween the groups ,all P>0 .05 .Compared with the patients in non‐bundle therapy group ,the APACHE‐Ⅱscore and SOFA score in 1 ,3 ,7 d after treatment significantly decreased in bundle therapy group ,all P<0 .05 .mortality rate in bundle therapy group and non‐bundle therapy group were 15 .38% and 35 .71% ,respectively ,P<0 .05 ;and length of ICU were(9 .04 ± 4 .48)d and(7 .00 ± 2 .32)d ,respectively ,P<0 .05 .Conclusion Early bundle therapy can effectively alleviate the severity of the disease and reduce mor‐tality of patients with septic shock after PCNL .