玉米直链淀粉生物合成多基因转化载体构建

淀粉分支酶基因sbe是影响玉米直链淀粉含量的主要因素,淀粉分支酶分为3种即sbeI、sbeIIa和sbeIIb,其中sbeIIb对直链淀粉含量影响效应最大,抑制玉米淀粉分支酶sbeIIb基因的表达可减少支链淀粉的含量,从而达到提高直链淀粉的目的;ADP-葡萄糖焦磷酸化酶(AGPase)是直链淀粉合成的关键酶,通过提高AGP表达量同样可提高玉米直链淀粉含量。以此为目的分别克隆了sbeIIb一段375bp高度保守区,玉米SBE基因一段175bp内含子,AGP完整开放阅读框,大麦胚乳特异启动子和ADPG基因终止子。构建了sbeIIb基因正、反义的hpRNA发夹结构,将该发夹结构与上述基因分别连接到pCAM-BIA3301上;构建得到包含sbeIIb基因干扰结构与AGP基因过表达的pCAMB-RSA多基因胚乳特异表达载体。为此,pCAMB-RSA载体的成功构建将为高直链淀粉玉米的培育奠定基础。     

Diagnostic Evaluation of Des-Gamma-Carboxy Prothrombin versus α-Fetoprotein for Hepatitis B Virus-Related Hepatocellular Carcinoma in China: A Large-Scale,Multicentre Study

An efficient serum marker for hepatocellular carcinoma (HCC) is currently lacking and requires intensive exploration. We aimed to evaluate the performance of des-gamma-carboxy prothrombin (DCP) for identifying hepatitis B virus-related HCC in a large, multicentre study in China. A total of 1034 subjects in three cohorts (A, B, and C) including HCC and various non-HCC controls were enrolled from 4 academic medical centers in China from January 2011 to February 2014. Blind parallel detections were conducted for DCP and AFP. The area under the receiver operating characteristic curve (AUC) was used to evaluate the diagnostic efficacies. In cohort A, which comprised 521 subjects, including patients with HCC, liver metastasis, liver cirrhosis (LC), and liver hemangiomas as well as healthy controls (HCs), the accuracy of DCP for distinguishing HCC from various controls was 6.2–9.7% higher than that of AFP. In cohort B, which comprised 447 subjects, including patients with HCC, LC, and chronic hepatitis B as well as HC, the accuracy of DCP was further elevated (12.3–20.67% higher than that of AFP). The superiority of DCP to AFP was more profound in the surveillance of early HCC [AUC 0.837 (95% CI: 0.771–0.903) vs. 0.650 (0.555–0.745)] and AFP-negative HCC [AUC: 0.856 (0.798–0.914)] and in discriminating HCC from LC (accuracy: 92.9% vs.64.71%). Higher DCP levels were associated with worse clinical behaviors and shorter disease-free survival. DCP not only is complementary to AFP in identifying AFP-negative HCC and in excluding AFP-positive non-HCC (liver cirrhosis), but also demonstrates improved performance in HCC surveillance, early diagnosis, treatment response and recurrence monitoring in the HBV-related population.     

活性部位的柔性

比较酶在变性过程中构象和活力变化,发现在活性完全丧失时尚无可察 觉的整体构象变化。排除变性剂抑制和寡聚酶解聚等可能性之后,提出了酶活性部位柔性假说。随后用多种实验方法直接证实了活性部位的构象变化先于分子整体构象变化,并与活性丧失同步,根据催化过程中活性部位构旬变化,以及限制活性部位构象变化对酶活性的影响,提出了酶活性部位柔性为酶充分表现其催化活性所必需的设想。     

Cofactor and CO2 donor regulation involved in reductive routes for polymalic acid production by Aureobasidium pullulans CCTCC M2012223

Polymalic acid (PMA) is a water-soluble polyester with many attractive properties for biomedical application. Its monomer l-malic acid is widely used in the food industry and also a potential C4 platform chemical. Cofactor and CO₂ donor involved in the reductive routes were investigated for PMA production by Aureobasidium pullulans. Biotin as the key cofactor of pyruvate carboxylase was favor for the PMA biosynthesis. Na₂CO₃ as CO₂ donor can obviously improved PMA titer when compared with no CO₂ supplier NaOH, and also exhibit more advantages than the other donor CaCO₃ because of its water-soluble characteristic. A combinational process with addition of biotin 70 mg/L and Na₂CO₃ as the CO₂ donor was scaled-up in 50 L fermentor, achieving the high product 34.3 g/L of PMA and productivity of 0.41 g/L h. This process provides an efficient and economical way for PMA and malic acid production, and is promising for industrial application.     

A strategy for fusion expression and preparation of functional glucagon-like peptide-1 (GLP-1) analogue by introducing an enterokinase cleavage site

KGLP-1, a 31-amino acid glucagon-like peptide-1 (GLP-1) analogue, has a great therapeutic potential for anti-diabetes. In this work, a strategy for expression and purification of functional KGLP-1 peptide has been established. KGLP-1 cDNA was fused with glutathione S-transferase (GST), with an enterokinase cleavage site in the fusion junction. The recombinant fusion protein GST–KGLP-1 was affinity purified via the GST-tag, and then digested with enterokinase. The resulting GST part as well as the enzymes were eliminated by ultra-filtration followed by size exclusion chromatograph. The yield of purified KGLP-1 was approximately 12.1 mg/L, with purity of 96.18 %. The recombinant KGLP-1 was shown to have similar bioactivity as native GLP-1 when evaluated in a Chinese hamster ovary cell line expressing a GLP-1 receptor-egfp reporter gene.     

植物微核技术在监测水质污染中的应用

应用韭菜微核技术,检测哈尔滨市量具刃具厂含铬废水的遗传毒性,试验表明：含铬废水可诱导韭菜和蚕豆根尖细胞产生微核,进行回归分析,其蚕豆对Ｃｒ＾＋６诱变剂回归分析为ｙ＝２４．９４＋１２７．７７ｘ（ｒ＝０．９９６,Ｐ〈０．０１）,韭菜对Ｃｒ＾＋６诱变剂回归广泛为：ｙ＝２１．７４＋１００．２３ｘ（ｒ＝０．９００,Ｐ〈０．０１）,ｒ＝１１．１７＋３．３８ｘ（ｒ＝０．７８１,Ｐ〈０．０５）,韭菜根尖微核技术具     

发酵性丝孢酵母HWZ004利用水稻秸秆水解液发酵产油脂

为高效利用水稻秸秆中的纤维素和半纤维素产油脂,采用稀酸预处理和酶水解两步法对水稻秸秆进行水解,然后以水解液为碳源,培养发酵性丝孢酵母Trichosporon fermentans HWZ004产微生物油脂。结果表明,经简单overliming法脱毒后水稻秸秆水解液中乙酸、糠醛和5-羟甲基糠醛的浓度分别为0.4 g/L、0.1 g/L和0.05 g/L。只需添加少量氮源和微量CuSO4?5H2O,该水解液即可满足T. fermentans HWZ004发酵产油脂的要求。发酵最适接种量、初始pH和温度分别是5.0％、7.0和25 ℃。T. fermentans HWZ004在优化条件下培养7 d的生物量、油脂含量和油脂产量分别是26.4 g/L,52.2％和13.8 g/L;油脂得率系数为17.0,大大高于驯化前菌株T. fermentans CICC 1368在脱毒水稻秸秆半纤维素水解液中的对应值 (11.9)。所产油脂的脂肪酸组成与植物油相似,不饱和脂肪酸含量达70％以上,宜作为生物柴油的生产原料。     

洞察景观环境影响蜜蜂之新视角: 肠道微生物

作为生态服务提供者的传粉蜜蜂与景观生态息息相关, 而以农田为主的景观组成显著降低了传粉蜜蜂的多样性。目前调查研究显示, 农田的扩张与蜜蜂多样性下降相关, 且农药残留对蜜蜂损害严重。景观中的开花植物决定了蜜蜂的食物(营养)组成, 其中花粉蛋白含量与蜜蜂的生长发育紧密相关。尽管研究已证实景观环境会显著影响蜜蜂蜂群的发展和个体的生长繁殖能力, 但未来还需要加强景观组成变化直接作用于蜜蜂的机制研究。另一方面, 大量研究表明蜜蜂肠道共生菌是影响宿主健康的重要因素: 可促进宿主吸收营养和抵抗病原菌。作为传粉者, 蜜蜂接触到的主要外部环境——花粉和花蜜都含有特殊的微生物, 很多研究暗示花源微生物是蜜蜂肠道菌来源之一。研究表明景观环境相关的食物(营养)、农药残留以及环境微生物都会显著影响肠道微生物。现有少量的研究证明不同景观的蜜蜂肠道微生物有差异, 景观环境可能通过作用于蜜蜂肠道微生物进而影响蜜蜂健康。然而不同景观环境中的微生物, 尤其是花源微生物和蜜蜂肠道菌之间的关联有待证明。景观对蜜蜂肠道微生物的影响值得研究, 希望可以从肠道菌的视角鉴别对蜜蜂友好的景观环境, 进而指导土地合理利用和蜜蜂保护。     

用一种改进的电泳方法测定抗冻蛋白的分子量

针对抗冻蛋白分子量较小的特点,该文采用了一种改进的Ｔｒｉｃｉｎｅ－ＳＤＳ－ＰＡＧＥ法测定其分子量。采用三层胶系统并在电极缓冲液中以Ｔｒｉｃｉｎｅ代替Ｇｌｙｃｉｎｅ。