Heterotetramers formed by an S-layer-streptavidin fusion protein and core-streptavidin as a nanoarrayed template for biochip development

Based on the S-layer protein SbpA of Bacillus sphaericus CCM 2177, an S-layer-streptavidin fusion protein was constructed. After heterologous expression, isolation of the fusion protein, and refolding, functional heterotetramers were obtained that had retained the ability to recrystallize into the square-lattice structure on plain gold chips and on gold chips precoated with secondary cell wall polymer (SCWP), which is the natural anchoring molecule for the S-layer protein in the bacterial cell wall. Monolayers generated by recrystallization of heterotetramers on plain gold chips or on gold chips precoated with thiolated SCWP were exploited for the binding of biotinylated oligonucleotides (30-mers). Hybridization experiments with complementary fluorescently labeled oligonucleotides carrying one mismatch or no mismatch (both 15-mers) were performed and evaluated with surface-plasmon-field-enhanced fluorescence spectroscopy. For surfaces generated by the recrystallization of heterotetramers on SCWP-coated gold chips, a detection limit of 1.57 pM could be determined, whereas for surfaces obtained by direct recrystallization of heterotetramers on plain gold chips, a detection limit of 8.2 pM was found. Measuring the association and dissociation processes of oligonucleotides carrying no mismatch led to a dissociation constant of K(D)=6.3 x 10(-10) m, whereas for oligonucleotides carrying one mismatch a dissociation constant of K(D)=7.9 x 10(-9) m was determined. This finding was confirmed by measuring the whole Langmuir isotherm, which resulted in a dissociation constant of K(D)=2.6 x 10(-8) m.     

超声波对荧光原位杂交技术检测氨氧化菌的影响

介绍了荧光原位杂交技术的基本原理和操作步骤。在采用荧光原位杂交技术检测氨氧化菌时必须经过预处理,而超声波对细胞的破碎是预处理中的关键步骤。试验通过比较预处理中超声波的三个强度对荧光原位杂交技术检测SBR反应器中氨氧化菌的效果,筛选出最佳的细胞破碎强度。结果显示,经过强度为150W超声破碎后,在荧光显微镜下观察,无论是丰度还是亮度方面,效果均比经过强度为100W或200W破碎后的好。     

Distinguishing activity decay and cell death from bacterial decay for two types of methanogens

As bacterial decay consists of cell death and activity decay, and the corresponding information about AOB/NOB, OHO, PAOs and GAOs has been experimentally acquired, another functional type of bacteria in biological wastewater treatment, methanogens, remains to be investigated, to gather the same information, which is extremely important for such bacteria with low growth rates. With successfully selection and enrichment of both aceticlastic and hydrogenotrophic methanogens, and by means of measuring specific methane activity (SMA) and hydrogen consumption rate (HCR), a series of decay experiments and molecular techniques such as FISH verification and LIVE/DEAD staining revealed, identified and calculated the decay and death rates of both aceticlastic and hydrogenotrophic methanogens respectively. The results indicated that the decay rates of aceticlastic and hydrogenotrophic methanogens were 0.070 and 0.034 d⁻¹ respectively, and the death rates were thus calculated at 0.022 and 0.016 d⁻¹ respectively. For this reason, cell deaths were only responsible for 31% and 47% of the total bacterial decay of aceticlastic and hydrogenotrophic methanogens, and activity decays actually contributed significantly to the total bacterial decay, respectively at 69% and 53%.     

通过油菜与播娘蒿远缘杂交获得黄籽油菜种质

对双低甘蓝型油菜（NJ5280）与播娘蒿远缘杂种自交后代进行选择和品质鉴定,在F3群体中得到13份黄籽材料,其中有4份黄籽双低高油油菜新种质,其品质符合含油量大于45％、硫苷低于30μmol／g、芥酸小于0．5％的双低高油标准。在F4群体中又得到24份黄籽材料,其中有8份黄籽双低高油甘蓝型油菜新种质。因此,通过甘蓝型油菜与播娘蒿远缘杂种后代的选择,可获得黄籽双低高油油菜优异种质。     

儿童侵袭性成熟B细胞淋巴瘤的临床病理学及分子遗传学特点

目的 分析总结中国儿童各类型侵袭性成熟B细胞淋巴瘤的临床病理学及分子遗传学特点,为其诊断的标准化提供依据.方法 收集97例儿童侵袭性成熟B细胞淋巴瘤石蜡包埋组织标本,包括伯基特淋巴瘤(BL)81例、弥漫大B细胞淋巴瘤(DLBCL)8例、介于BL和DLBCL间的不能分类的B细胞淋巴瘤(BL/DLBCL)8例,利用免疫组织化学技术和间期荧光原位杂交(FISH)技术检测其免疫表型和分子遗传学特征.结果 BL的bcl-2和MUM1的阳性率分别为3%(2/66)和17%(12/71),DLBCL分别为50%(4/8)和63%(5/8),BL/DLBCL分别为50%(4/8)和63%(5/8).BL、DLBCL和BL/DLBCL的Ki-67平均值分别为(93±4.4)%、(83±14.3)%和(80±11.5)%.BL、DLBCL和BL/DLBCL的c-myc基因易位的比例分别为98%(79/81)、38%(3/8)和50%(4/8).38%(3/8)的DLBCL和25%(2/8)的BL/DLBCL存在bcl-6基因的多拷贝,BL与DLBCL之间、BL与BL/DLBCL之间bcl-2、MUM1和Ki-67平均值的差异及c-myc基因易位和bcl-6基因多拷贝的差异均有统计学意义(均P＜0.05).结论 儿童侵袭性成熟B细胞淋巴瘤的诊断和分型需要综合分析形态学、免疫表型和分子遗传学特征.儿童BL/DLBCL可能是DLBCL的一个亚型.CD10+、bcl-6+、bcl-2-、Ki-67＞90%、伴有IGH/c-myc重排、不伴有bcl-2和bcl-6重排时,支持BL的诊断;bcl-2+、Ki-67为50%～90%,同时伴有bcl-6基因的多拷贝时,支持DLBCL或BL/DLBCL的诊断.     

金纳米球壳结构局域表面等离子体共振调谐特性

理论研究了金纳米球壳结构局域表面等离子体共振(LSPR)的调谐特性。结果表明,对于固定内径的球壳结构,利用杂化效应使得球壳结构可通过改变其厚度实现LSPR峰位的调控,并可保持较窄的谱宽;而对于不同内径的球壳,在LSPR调控过程中,随壳层厚度与球壳内半径比值的减小,吸收光谱在消光光谱中所占比例增大。不仅如此,利用杂化效应进行偶极峰位调控时所能获得的峰位调控范围与球壳内径有关,内径越大,所能获得的偶极峰位调控范围越大。     

Characterization of ten H2 producing cyanobacteria isolated from the Baltic Sea and Finnish lakes

The genetic background and activities of the enzymes involved in H₂ production were investigated from ten distinct H₂ producing cyanobacteria, revealed by a recent screening. All strains are N₂-fixing, filamentous and heterocystous. Southern hybridization revealed that the tested strains possess the genes encoding the conventional nitrogenase (nifHDK1), and lack the alternative nitrogenases. The high H₂ production rate of these strains was shown not to be dependent on the presence of highly active nitrogenase or bidirectional hydrogenase enzymes. Moreover, most of the strains possessed a highly active uptake hydrogenase enzyme. We also examined the structure of the nif and hup operons encoding nitrogenase and uptake hydrogenase enzymes in the Calothrix 336/3 strain, the best H₂ producer in the screening. We concluded that the ability of the cyanobacteria to produce high levels of H₂ is not directly linked to the maximum capacities of the enzymes involved in H₂ production.     

Harnessing P450 Enzyme for Biotechnology and Synthetic Biology

Cytochrome P450 enzymes (P450s, CYPs) catalyze the oxidative transformation of a wide range of organic substrates. Their functions are crucial to xenobiotic metabolism and steroid transformation in humans and other organisms. The enzymes are promising for synthetic biology applications but limited by several drawbacks including low turnover rates, poor stability, the dependance of expensive cofactors and redox partners, and the narrow substrate scope. To conquer these obstacles, emerging strategies including substrate engineering, usage of decoy and decoy-based small molecules auxiliaries, designing of artificial enzyme cascades and the incorporation of materials have been explored based on the unique properties of P450s. These strategies can be applied to a wide range of P450s and can be combined with protein engineering to improve the enzymatic activities. This minireview will focus on some recent developments of these strategies which have been used to leverage P450 catalysis. Remaining challenges and future opportunities will also be discussed.