Genetic diversity of the threatened Chinese endemic plant,Sinowilsonia henryi Hemsi. (Hamamelidaceae), revealed by inter-simple sequence repeat (ISSR) markers

Sinowilsonia henryi Hemsi., the only representative of the monotypic genus Sinowilsonia Hemsi. (Hamamelidaceae), is a threatened plant endemic to China with high phylogenetical, ecological and economical values. In the present study, inter-simple sequence repeat (ISSR) markers were employed to investigate the genetic diversity and differentiation of 214 individuals sampled from 14 populations. Fifteen selected primers yielded a total of 178 bright and discernible bands. The genetic diversity was low at the population level (h = 0.1025; I = 0.1506; PPL = 26.7%), but quite high at the species level (h = 0.2449; I = 0.3690; PPL = 72.5%). In line with the limited gene flow (N_m = 0.3537), the hierarchical analysis of molecular variance (AMOVA) revealed pronounced genetic differentiation among populations (Φ_ST = 0.6639). Furthermore, the Mantel test revealed a significant correlation between genetic and geographic distances among populations (r = 0.688, P = 0.001), indicating the role of geographic isolation in shaping its present population genetic structure. The present patterns of genetic diversity of S. henryi were assumed to result largely from its evolutionary history and geographic factors. Based on these findings, conservation strategies were proposed to preserve this threatened plant.     

Membrane active antimicrobial activity and molecular dynamics study of a novel cationic antimicrobial peptide polybia-MPI,from the venom of Polybia paulista

As the frequent emergence of the resistant bacteria, the development of new agents with a new action mode attracts a great deal of interest. It is now widely accepted that antimicrobial peptides (AMPs) are promising alternatives to conventional antibiotics. In this study, antimicrobial peptide polybia-MPI and its analogs were synthesized and their antibacterial activity was studied. Our results revealed that polybia-MPI has potent antibacterial activity against both Gram-positive and Gram-negative bacteria. Its ability to make PI permeate into bacteria and lead to the leakage of calcein from model membrane LUVs, suggests a killing mechanism involving membrane perturbation. SEM and TEM microscopy experiments verified that the morphology of bacteria was changed greatly under the treatment of polybia-MPI. Compared with the conventional chemotherapy, polybia-MPI targets the cell membrane rather than entering into the cell to exert its antibacterial activity. Furthermore, molecular dynamics (MD) simulations were employed to investigate the mechanism of membrane perturbation. The results indicated that the α-helical conformation in the membrane is required for the exhibition of antibacterial activity and the membrane disturbance by polybia-MPI is a cooperative process. In conclusion, with the increasing resistance to conventional antibiotics, there is no doubt that polybia-MPI could offer a new strategy to defend the resistant bacteria.     

Loss of second and sixth conserved cysteine residues from trypsin inhibitor-like cysteine-rich domain-type protease inhibitors in Bombyx mori may induce activity against microbial proteases

Previous studies have indicated that most trypsin inhibitor-like cysteine-rich domain (TIL)-type protease inhibitors, which contain a single TIL domain with ten conserved cysteines, inhibit cathepsin, trypsin, chymotrypsin, or elastase. Our recent findings suggest that Cys^2nd and Cys^6th were lost from the TIL domain of the fungal-resistance factors in Bombyx mori, BmSPI38 and BmSPI39, which inhibit microbial proteases and the germination of Beauveria bassiana conidia. To reveal the significance of these two missing cysteines in relation to the structure and function of TIL-type protease inhibitors in B. mori, cysteines were introduced at these two positions (D36 and L56 in BmSPI38, D38 and L58 in BmSPI39) by site-directed mutagenesis. The homology structure model of TIL domain of the wild-type and mutated form of BmSPI39 showed that two cysteine mutations may cause incorrect disulfide bond formation of B. mori TIL-type protease inhibitors. The results of Far-UV circular dichroism (CD) spectra indicated that both the wild-type and mutated form of BmSPI39 harbored predominantly random coil structures, and had slightly different secondary structure compositions. SDS-PAGE and Western blotting analysis showed that cysteine mutations affected the multimerization states and electrophoretic mobility of BmSPI38 and BmSPI39. Activity staining and protease inhibition assays showed that the introduction of cysteine mutations dramaticly reduced the activity of inhibitors against microbial proteases, such as subtilisin A from Bacillus licheniformis, protease K from Engyodontium album, protease from Aspergillus melleus. We also systematically analyzed the key residue sites, which may greatly influence the specificity and potency of TIL-type protease inhibitors. We found that the two missing cysteines in B. mori TIL-type protease inhibitors might be crucial for their inhibitory activities against microbial proteases. The genetic engineering of TIL-type protease inhibitors may be applied in both health care and agricultural industries, and could lead to new methods for breeding fungus-resistant transgenic crops and antifungal transgenic silkworm strains.     

铁扫帚中总多糖提取工艺的研究

研究了铁扫帚中总多糖提取的最佳工艺,并对其进行了薄层层析,结果表明:A3B2C1条件下,总多糖提取率最高。薄层层析得两个组分,其Rf值分别为:0.65和0.55     

植物对氨基酸的吸收研究进展

氨基酸在提高植物产量、改善产品品质、增强植株抗逆性、保护生态环境等方面发挥着越来越重要的作用,在农业生产中越来越受到重视。本文简述了氨基酸含量、氨基酸种类和植物种类对植物吸收氨基酸的影响,并对氨基酸营养研究进行展望,以期提高人们对植物氨基酸营养的认识,促进氨基酸在农业中的应用和发展。     

大孔树脂分离纯化“黑金刚”紫马铃薯花青苷工艺研究

以紫色马铃薯"黑金刚"花青苷为原料,采用D101、HDP100A、HDP450A、NK-9、AB-8五种大孔吸附树脂对花青苷的吸附与解析特性进行了比较研究,并在此基础上,采用最佳大孔树脂对花青苷纯化过程中的静态、动态吸附和解析附条件进行了优化研究。结果表明AB-8大孔树脂具有较好的吸附和解析能力,是纯化紫色马铃薯花青苷的最佳树脂,较优纯化条件为:上样液花青苷浓度为0.028mg.g-1,上样液pH=2,洗脱液乙醇浓度为50%,洗脱液pH=1,吸附流速为1mL.min-1,洗脱流速为1mL.min-1。经大孔树脂纯化后,色价值比纯化前提高了7.55倍。     

Isolation and characterization of endophytic fungi from Camptotheca acuminata

In this study, a total of 161 endophytic fungal isolates from Camptotheca acuminata were obtained and classified to 16 taxa according to morphological and molecular analysis. These taxa were composed of 2 frequent genera (Botryosphaeria and Fusarium) and 14 infrequent groups such as Xylaria, Diaporthe, Rhizopus, Epicoccum, and Preussia, demonstrating that fungal endophytes in C. acuminata were highly abundant and diverse. Antimicrobial activity screening using filter-paper diffusion method showed that 47.6 % of the tested isolates had antimicrobial activity against at least one of the test microorganisms. Screening of fungal endophyte-derived camptothecin analogues by TLC and LC–MS/MS³ demonstrated that a strain Botryosphaeria dothidea, X-4 could produce 9-methoxycamptothecin (9-MCPT) when cultured in Sabouraud’s dextrose broth for 12 days under shake flask and bench-scale fermention conditions. This work showed that the fungal endophytes from C. acuminata could be an alternative source for the production of 9-MCPT and other natural antimicrobial compounds.     

蒲公英植酸对沙门氏菌抑制作用及其抑菌机理研究

为探究蒲公英植酸对沙门氏菌的抑制作用及其抑菌机理。本文利用沉淀法和离子交换法提取蒲公英植酸,滤纸片法分析蒲公英植酸对沙门氏菌(Salmonella)的抑菌作用,倍比稀释法研究蒲公英植酸的最低抑菌浓度。通过分析沙门氏菌的细胞通透性和生长动力学,结合扫描电镜和荧光显微镜研究了蒲公英植酸对沙门氏菌的抑菌机理,表明蒲公英植酸对沙门氏菌具有很好的抑菌能力,其最小抑菌浓度为0.2 mg/mL。而且植酸对沙门氏菌的抑制作用是通过破坏细胞膜达到抑菌的效果,并且植酸浓度越高,抑菌效果越显著。这表明蒲公英植酸可以有效地抑制沙门氏菌生长,其主要是通过破坏菌体细胞膜完整性,增加细胞薄膜的通透性,使细胞内容物外溢达到抑制细菌生长的目的。     

大鲵皮肤色素提取工艺及抗氧化研究

为优化大鲵皮肤黑色素的提取工艺条件,探讨大鲵皮肤黑色素组成成分及体外抗氧化活性,采用酶法和碱溶酸沉法提取大鲵皮肤黑色素,以氢氧化钠浓度、液料比、提取温度为影响色素提取率因素,优化黑色素提取工艺条件,用紫外-可见光谱仪、红外光谱仪和超高效液相质谱仪测定黑色素的光谱特性,测定其抗氧化性。结果表明:大鲵皮肤黑色素最佳提取工艺条件为氢氧化钠浓度1.5 mol/L、液料比1∶15、提取温度45℃,黑色素提取率达0.65%。大鲵皮肤黑色素的紫外最大吸收波长为214 nm,由真黑色素和脱黑色素两种色素组成,其对超氧阴离子自由基的清除率为30.51%,对羟基自由基的清除率为54.17%。大鲵皮肤黑色素具有一定的体外抗氧化能力。