<Emphasis Type="Italic">SQUA</Emphasis>-like genes in the orchid <Emphasis Type="Italic">Phalaenopsis</Emphasis> are expressed in both vegetative and reproductive tissues

The SQUA family (AP1/FUL family) of MADS-box genes plays an important role in the transition from the vegetative to the reproductive development of angiosperms, and its origin might be concurrent with fixation of floral structure in angiosperms. Here, we isolated two Phalaenopsis MADS-box genes designated ORAP11 and ORAP13, both of which belong to the monocot FUL-like clade of the SQUA family. RT-PCR showed that both genes are strongly expressed in the floral bud, and also detected in the vegetative organs. During later stages, ORAP11 was only detected in the column, but ORAP13 signal was absent from all of the floral organs. In-situ hybridization experiments detected both genes in the tips and margins of developing petals and lips, the developing column, and ovule. Over-expression of both genes in tobacco induced early flowering and changed plant architecture. Our results suggest that in Phalaenopsis, both genes might share partly redundant activities and play important roles in the process of floral transition and morphological architecture. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Griffithsin, a potent HIV entry inhibitor, is an excellent candidate for anti-HIV microbicide

BACKGROUND: The predominant mode of HIV-1 transmission is by heterosexual contact. The cervical/vaginal mucosa is the main port of HIV entry in women. A safe and effective topical microbicide against HIV is urgently needed to prevent sexual transmission. Hence, we evaluated griffithsin (GRFT), a 12.7 kDa carbohydrate-binding protein, both native and recombinant GRFT, potently inhibited both CXCR4-and CCR5-tropic HIV infection and transmission in vitro. METHODS: The antiviral efficacy of native and recombinant GRFT against CXCR4-and CCR5-tropic HIV and SHIV strains and SIVmac251 was evaluated by in vitro assays. We also evaluated the time course of antiviral activity and stability of GRFT in cervical/vaginal lavage as a function of pH 4-8. RESULTS: Griffithsin blocked CXCR4-and CCR5-tropic viruses at less than 1 nm concentrations and exhibited a high potency. GRFT was stable in cervical/vaginal lavage fluid and maintained a similar potency of anti-HIV activity. GRFT is not only a highly potent HIV entry inhibitor, but also prevents cell fusion and cell-to-cell transmission of HIV. CONCLUSIONS: The in vitro efficacy of GRFT revealed low cytotoxicity, high potency, rapid onset of antiviral activity and long-term stability in cervical/vaginal lavage. GRFT is an excellent candidate for anti-HIV microbicide development.     

Phenotypes and fed-batch fermentation of ubiquinone-overproducing fission yeast using ppt1 gene

Ubiquinone (UQ), a component of the electron transfer system in many organisms, has been widely used for pharmaceuticals and cosmetics. In this study, we cloned and overexpressed the full-length ppt1 (MTppt1) gene, which encodes p-hydroxybenzoate:polyprenyltransferase and ERppt1 gene, which was modified to be localized on endoplasmic reticulum in fission yeast. The yeast MTppt1 and ERppt1 transgenic lines showed about 3.7 and 5.1 times increment in UQ content and the recombinant yeasts with a higher UQ level are more resistant to H(2)O(2), Cu(2+) and NaCl, and interestingly their growth was also faster than the wild type at lower temperature. For large-scale cultivation, the direct feedback control of glucose using an on-line ethanol concentration monitor for ubiquinone production of yeast ERppt1 by high-cell-density fermentation was investigated and the fermentation parameters (e.g., dissolved oxygen, pH, ethanol concentration, oxygen uptake rate, carbon dioxide evolution rate and respiration quotient) were also discussed. After 90 h cultures, the yeast dry cell weight reached 57 gl(-1) and the ubiquinone yield reached 23 mgl(-1). In addition, plasmid stability was maintained at high level throughout the fermentation.     

Fermentation of 1,3-propanediol by a lactate deficient mutant of Klebsiella oxytoca under microaerobic conditions

Klebsiella oxytoca M5al is an excellent 1,3-propanediol (1,3-PD) producer, but too much lactic acid yielded greatly lessened the fermentation efficiency for 1,3-PD. To counteract the disadvantage, four lactate deficient mutants were obtained by knocking out the ldhA gene of lactate dehydrogenase (LDH) of K. oxytoca M5al. The LDH activities of the four mutants were from 3.85 to 6.92% of the parental strain. The fed-batch fermentation of 1,3-PD by mutant LDH3, whose LDH activity is the lowest, was studied. The results showed that higher 1,3-PD concentration, productivity, and molar conversion rate from glycerol to 1,3-PD can be gained than those of the wild type strain and no lactic acid is produced under both anaerobic and microaerobic conditions. Sucrose fed during the fermentation increased the conversion and sucrose added at the beginning increased the productivity. In fed-batch fermentation with sucrose as cosubstrate under microaerobic conditions, the 1,3-PD concentration, conversion, and productivity were improved significantly to 83.56 g l⁻¹, 0.62 mol mol⁻¹, and 1.61 g l⁻¹ h⁻¹, respectively. Furthermore, 60.11 g l⁻¹ 2,3-butanediol was also formed as major byproduct in the broth.     

中国水仙的核型分析和小孢子发生中的细胞学研究

中国水仙(Narcissustazettavar.chinensis)只开花不结实,以鳞茎营养繁殖。对中国水仙的染色体倍性有不同的报道。对中国水仙的核型分析,支持它是三倍体的观点,但其核型也显示出了异源三倍体的倾向。在小孢子母细胞减数分裂过程中,染色体的异常行为多表现为:中期出现单个染色体游离在纺锤体外面、在后期出现的染色体桥和落后染色体、在末期出现的单个染色体游离在细胞核外形成微核的现象。这些异常现象引起小孢子的败育,也支持中国水仙为三倍体植物的观点。     

Plant biodiversity and its conservation strategy in the in-undation and resettlement districts of the Yangtze Three Gorges, China

The investigation on and conservation project of plant biodiversity in the inundation and resettlement districts of the Three Gorges region were carried out with the ongoing construction of the Three Gorges Dam. Based on the first-hand specimens collected for many years and the historical records, we found that there were about 1784 higher plant species, belonging to 762 genera of 170 families, which accounted for 59.19%, 75.30% and 85.85% of the total amount of species, genera and families in the Three Gorges region, including 27 species specific in these two districts. The shrub and grass communities were mostly distributed along the low-altitude river bank zone of the Yangtze River and were largely influenced by the dynamic water level of the Three Gorges reservoir area. The measurement of Shannon-Wiener and Pielou evenness indexes on 21 Pinus massoniana (Masson pine) communities distributed in the Three Gorges reservoir area was carried out, and results showed that the biodiversity of shrub layers was the highest, followed by grass layers, the middle, and tree layers, the lowest. Among the 11 typical shrub communities in this region, the abundance of shrub layers of Form. Rhus chinensis, Form. Cotinus coggygria var. cinerea, Form. Vitex negundo var. cannabifoliaand and Form. Coriaria sinica was relatively higher, which were 16, 26, 29 and 15, respectively. These four shrubs also showed high biodiversity index (H′), which were 1.791, 3.427, 2.949 and 1.718, respectively. Among the 9 grasslands mainly distributed along the river bank zone, 4 grassland communities with higher altitude (Form. Dicranopteris dichotoma, Form. Miscanthus floridutus, Form. Imperata cylindrica var. major and Form. Arthraxon hispidus) showed higher abundance and biodiversity, which were 1.697, 1.354, 1.144 and 1.018, respectively. Considering the inundation and resettlement districts, there were 22 vegetation types threatened by submersion, including 4 woody plant communities, 9 shrub communities and 9 grassland communities. The 6 shrub communities of Form. Securinega wuxiensis, Form. Bauhinia brachycarpa, Form. Geum aleppicum, Form. Distylium chinense, Form. Buxus henryi and Form. Myrsine africana would be fully submerged and part of Adianthum reniforme var. sinensis and Buxus ichangensis would be also submerged. At present, plant species conservation and monitoring stations in the Three Gorges region have been constructed, and more than 200 plant species have already been transplanted and conserved, including 37 threatened species listed in the China Plant Red Data Book and 11 local constructive species.     

Synthesis and biological evaluation of nitric oxide-releasing derivatives of oleanolic acid as inhibitors of HepG2 cell apoptosis

A total of 106 nitric oxide-releasing derivatives of oleanolic acid were synthesized and their effects on the inhibition of anti-Fas-mediated HepG2 cell apoptosis were evaluated in vitro. Several compounds inhibited anti-Fas-mediated HepG2 cell apoptosis in a dose-dependent manner. Within this series of compounds, 8b is the most potent inhibitor. The development of new NO-releasing derivatives of oleanolic acid may aid in the design of NO-based medicines for the intervention of human liver inflammatory diseases.     

Structure-based design of robust glucose biosensors using a Thermotoga maritima periplasmic glucose-binding protein

We report the design and engineering of a robust, reagentless fluorescent glucose biosensor based on the periplasmic glucose-binding protein obtained from Thermotoga maritima (tmGBP). The gene for this protein was cloned from genomic DNA and overexpressed in Escherichia coli, the identity of its cognate sugar was confirmed, ligand binding was studied, and the structure of its glucose complex was solved to 1.7 Angstrom resolution by X-ray crystallography. TmGBP is specific for glucose and exhibits high thermostability (midpoint of thermal denaturation is 119 +/- 1 degrees C and 144 +/- 2 degrees C in the absence and presence of 1 mM glucose, respectively). A series of fluorescent conjugates was constructed by coupling single, environmentally sensitive fluorophores to unique cysteines introduced by site-specific mutagenesis at positions predicted to be responsive to ligand-induced conformational changes based on the structure. These conjugates were screened to identify engineered tmGBPs that function as reagentless fluorescent glucose biosensors. The Y13C*Cy5 conjugate is bright, gives a large response to glucose over concentration ranges appropriate for in vivo monitoring of blood glucose levels (1-30 mM), and can be immobilized in an orientation-specific manner in microtiter plates to give a reversible response to glucose. The immobilized protein retains its response after long-term storage at room temperature.     

Role of follicle stimulating hormone and epidermal growth factor in the development of porcine preantral follicle in vitro

The aim of the present study was to assess the role of follicle stimulating hormone (FSH), epidermal growth factor (EGF) or a combination of EGF and FSH on the in vitro growth of porcine preantral follicles, estradiol secretion, antrum formation, oocyte maturation and subsequent embryonic development. Porcine preantral follicles were cultured for 3 days in the absence or in the presence of FSH or EGF. Oocytes from these follicles were then matured, fertilized in vitro and embryos were cultured. Estradiol secretion and histological analysis of cultured follicles were also carried out. The results showed that when FSH, or a combination of EGF and FSH, was added to the culture medium, most of preantral follicles grew to antral follicles with high estradiol secretion and the oocytes from these antral follicles could mature, fertilize and develop to the blastocyst stage. Without FSH, or a combination of EGF and FSH, preantral follicles were unable to develop to the antral stage. Histology demonstrated that the resulting follicles were nonantral, estradiol production was reduced and none of their oocytes matured after in vitro maturation. The results indicate the essential role of FSH in promoting in vitro growth of porcine preantral follicle, estradiol secretion, antrum formation, oocyte maturation and subsequent embryonic development. EGF with FSH treatment of porcine preantral follicles improves the quality of oocytes, shown by a higher frequency of embryonic development.     

东海原甲藻(Prorocentrum donghaiensis)对原生动物群落结构影响的现场船基实验

2005年在长江口赤潮频发海域调查期间,分别于4月27日、5月4日和5月8日,在zzf1、zc18a和ra5站位利用现场船基培养的方法,研究了添加到赤潮密度10^6cells L^-1的东海原甲藻（Prorocentrum donghaiense）对原生动物群落结构的影响。结果发现,赤潮密度的东海原甲藻抑制了小型无壳纤毛虫的种群数量,而没有抑制中大型砂壳纤毛虫和夜光虫（Noctiluca scintillans）的种群数量,从而使得原生动物群落向中大型种类演替,这种影响的程度与原生动物本身的群落组成和浮游植物的组成密切相关。添加东海原甲藻72h后,在以小型无壳纤毛虫管游虫（Cyrtostrombidium sp．）为优势种的zzf1站位,演替为以大型原生动物夜光虫为优势种的群落;在以中大型原生动物百乐拟铃虫（Tintinnospsis beroidea）和夜光虫为优势种的zc18a站位,仍然是以此为优势种,且大型原生动物夜光虫在群落中的比例上升;在以小型无壳纤毛虫急游虫2（Strombidium sp．2）为优势种的ra5站位,演替为以中大型原生动物百乐拟铃虫和亚速岛网纹虫（Favella azorica）为优势种的群落。zzf1和ra5站位实验组中原生动物的总丰度都呈下降趋势,而zc18a站位变化不明显,这是由于前两个站位的最优势种管游虫（zzf1站位）和急游虫2（ra5站位）的丰度迅速下降,而zc18a站位的优势种百乐拟铃虫和夜光虫的丰度比较稳定造成的。在zzf1和zc18a站位,对照组和实验组中原生动物的总生物量在实验前后都没有大的变化,而在ra5站位却均呈下降的趋势。这可能与浮游植物的组成有关,zzf1和zc18a站位是以无毒的锥状斯氏藻（Scrippsiella trochoidea）和螺旋环沟藻（Gyrodinium spirale）为主,而ra5站位是以有毒的亚历山大藻（Alexandriumsp．）为主。综上可见,可预测当东海大规模赤潮爆发时,会使原生动物群落向中大型种类演替,进而可能影响海洋生态系统的结构和功能。