不同预处理方法影响昆虫干标本DNA提取效果

为从昆虫干标本中获取高质量DNA,以利于后期PCR扩增目的基因,本研究对DNA提取前昆虫干标本的预处理方法进行探讨,分别测定其纯度和质量,并用不同引物进行了扩增效果对比。结果显示:经过预处理的样品DNA平均含量均高于CK的52.283 ng/滋L,最高为0.9%Na Cl处理3 h,平均含量达122.632 ng/滋L;米象4种预处理方法提取的DNA利用引物Lco1490/Hco2198及UEA7/UEA10均能扩增出650 bp长度条带,仅有0.9%Na Cl溶液浸泡3 h处理的米象可通过引物J173/J1331扩增出1 000 bp的长片段条带。研究表明,经过预处理后,一定程度上减少了基因组DNA的断链,主要对样品DNA的质量浓度影响显著;以9%Na Cl溶液3 h预处理后的昆虫干标本,利用磁珠法所提DNA的质量及PCR扩增效果最佳。     

中国顶丝藻科（Acrochaetiaceae）新记录I

中国顶丝藻科新记录６种。密集旋体藻Ａｕｄｏｕｉｎｅｌｌｅｄｅｎｓａ,亮管旋体藻Ａ．ｈｙａｌｏｓｉｐｈｏｎｉａｅ,小旋体藻Ａ．ｐａｒｖｕｌａ,羽状旋体藻Ａ．ｐｌｕｍｏｓａ,顶生旋体藻Ａ．ｔｅｒｍｉｎａｌｉｓ,图氏旋体藻Ａ．ｔｈｕｒｅｔｉｉ。     

TSNAdb: A Database for Tumor-specific Neoantigens from Immunogenomics Data Analysis

Tumor-specific neoantigens have attracted much attention since they can be used as biomarkers to predict therapeutic effects of immune checkpoint blockade therapy and as potential targets for cancer immunotherapy. In this study, we developed a comprehensive tumor-specific neoantigen database (TSNAdb v1.0), based on pan-cancer immunogenomic analyses of somatic mutation data and human leukocyte antigen (HLA) allele information for 16 tumor types with 7748 tumor samples from The Cancer Genome Atlas (TCGA) and The Cancer Immunome Atlas (TCIA). We predicted binding affinities between mutant/wild-type peptides and HLA class I molecules by NetMHCpan v2.8/v4.0, and presented detailed information of 3,707,562/1,146,961 potential neoantigens generated by somatic mutations of all tumor samples. Moreover, we employed recurrent mutations in combination with highly frequent HLA alleles to predict potential shared neoantigens across tumor patients, which would facilitate the discovery of putative targets for neoantigen-based cancer immunotherapy. TSNAdb is freely available at http://biopharm.zju.edu.cn/tsnadb.     

中国河南两株庚型肝炎病毒（HGV）NS5区部分cDNA的克隆与序列分析

通过逆转录－聚合酶链反应从我国河南省２例重叠感染ＨＣＶ或ＨＢＶ／ＨＤＶ的献血啼中,分离到ＨＢＶＮＳ５区的部分ｃＤＮＡ,对其进行序列分析比较,结果表明,河南株ＨＧＶＮＳ５工核苷酸与两中国ＨＧＶ主同源性高于国外代表株（８８．５－９０．６％）,但由核苷酸推导的氨基酸的同源性都无明显的地区性区别。ＨＧＶＮＳ５区氨基酸序列较保守,缺乏明显高变区,中国４株ＨＧＶ在７３８４位发生了由Ｃ→Ｔ的变异,从而导致一个人     

Ricin A chain reaches the endoplasmic reticulum after endocytosis

Ricin is a potent ribosome inactivating protein and now has been widely used for synthesis of immunotoxins. To target ribosome in the mammalian cytosol, ricin must firstly retrograde transport from the endomembrane system to reach the endoplasmic reticulum (ER) where the ricin A chain (RTA) is recognized by ER components that facilitate its membrane translocation to the cytosol. In the study, the fusion gene of enhanced green fluorescent protein (EGFP)-RTA was expressed with the pET-28a (+) system in Escherichia coli under the control of a T7 promoter. The fusion protein showed a green fluorescence. The recombinant protein can be purified by metal chelated affinity chromatography on a column of NTA. The rabbit anti-GFP antibody can recognize the fusion protein of EGFP-RTA just like the EGFP protein. The cytotoxicity of EGFP-RTA and RTA was evaluated by the MTT assay in HeLa and HEP-G2 cells following fluid-phase endocytosis. The fusion protein had a similar cytotoxicity of RTA. After endocytosis, the subcellular location of the fusion protein can be observed with the laser scanning confocal microscopy and the immuno-gold labeling Electro Microscopy. This study provided important evidence by a visualized way to prove that RTA does reach the endoplasmic reticulum.     

5种光合细菌种间原生质体融合及优良农用融合子的筛选鉴定

处于对数生长期的光合细菌球形红假单胞菌(Rhodopseudomonassphaeroides)、沼泽红假单胞菌(Rhodopseudomonaspalustris)、嗜酸红假单胞菌(Rhodopseudomdnasacidophila)、深红红螺菌(Rhodospirarubrum)、万尼氏红微菌(Rhodomocrobiumvannielii),经溶菌酶(3mg/L)处理50min后,获得了它们的菌体形成的原生质体,其再生率分别为80%、71%、82%、61%、74%.取等量的亲本菌株在35%的PEG(MW6000)诱导下两两融合5min,共10种组合.其融合率为球×沼2.5×10-4、球×嗜2.1×10-4、球×深2.0×10-4、球×万2.1×10-4、沼×嗜2.8×10-4、沼×深2.4×10-4、沼×万2.6×10-4、嗜×深2.0×10-4、嗜×万2.3×10-4、深×万2.4×10-4.经影印法鉴定:形成的融合子可以分别生长于以相应的有机物为唯一碳源的培养基上,所有融合子体积均相当于两亲本株体积之和,融合子菌落形态特征介于两亲本株之间.从中随机挑选100个融合子,以辣椒苗作为靶标植物,从上述融合子中筛选到了1株具有显著促进作物生长、提高抗病性的融合子.     

Hdac2 regulates the cardiac hypertrophic response by modulating Gsk3 beta activity

In the adult heart, a variety of stresses induce re-expression of a fetal gene program in association with myocyte hypertrophy and heart failure. Here we show that histone deacetylase-2 (Hdac2) regulates expression of many fetal cardiac isoforms. Hdac2 deficiency or chemical histone deacetylase (HDAC) inhibition prevented the re-expression of fetal genes and attenuated cardiac hypertrophy in hearts exposed to hypertrophic stimuli. Resistance to hypertrophy was associated with increased expression of the gene encoding inositol polyphosphate-5-phosphatase f (Inpp5f) resulting in constitutive activation of glycogen synthase kinase 3beta (Gsk3beta) via inactivation of thymoma viral proto-oncogene (Akt) and 3-phosphoinositide-dependent protein kinase-1 (Pdk1). In contrast, Hdac2 transgenic mice had augmented hypertrophy associated with inactivated Gsk3beta. Chemical inhibition of activated Gsk3beta allowed Hdac2-deficient adults to become sensitive to hypertrophic stimulation. These results suggest that Hdac2 is an important molecular target of HDAC inhibitors in the heart and that Hdac2 and Gsk3beta are components of a regulatory pathway providing an attractive therapeutic target for the treatment of cardiac hypertrophy and heart failure.     

Fluorescent labeling for clonal selection of Marc 145 cells secreting high levels of recombinant protein PBD-1

Despite the powerful impact gene expression markers like the green fluorescent protein (GFP) or enhanced GFP (EGFP) exert on linking the expression of recombinant protein for selection of high producers in recent years, there is still a strong incentive to develop more economical and efficient methods for isolating mammalian cell clones secreting high levels of recombinant proteins. Here we present a new method based on the co-expression of EGFP that allows clonal selection in standard 96-well cell culture plates. The genes encoding the EGFP protein and the related protein are linked by an internal ribosome entry site and thus are transcribed into the same mRNA in an independent translation process. Since both proteins arise from a common mRNA, the EGFP expression level correlates with the expression level of the therapeutic protein in each clone. By expressing recombinant porcine β-defensin 1 in Marc 145 cells, we demonstrate the robustness and performance of this technique. The method can be served as an alternative to identify high-producer clones with various cell sorting methods.     

Fabrication of Bioinspired Structured Superhydrophobic and Superoleophilic Copper Mesh for Efficient Oil-water Separation

Oily water treatment has attracted the attention of many researchers.The development of effective and cheap oil/water separation materials is urgent for treating this problem.Herein,inspired by superhydrophobic typical plant leaves such as lotus,red rose and marigold,superhydrophobic and superoleophilic copper mesh was fabricated by etching and then surface modification with 1-dodecanethiol (HS(CH2)11CH3).A rough silver layer is formed on the mesh surface after immersion.The obtained mesh surface exhibits superhydrophobicity and superoleophilicity and the static water contact angle was 153° ± 3°.In addition,the as-prepared copper mesh shows self-cleaning character with water and chemical stability.The as-prepared copper foam can easily remove the organic solvents either on water or underwater.We demonstrate that by using the as-prepared mesh,oils can be absorbed and separated,and that high separation efficiencies of larger than 92％ are retained for various oils.Thus,such superhydrophobic and superoleophilic copper mesh is a very promising material for the application of oil spill cleanup and industrial oily wastewater treatment.