转Bt基因玉米根际土壤及秸秆残体中Cry1Ab蛋白含量动态

采用ELISA法研究了田间种植条件下转Bt基因玉米MON810生育期根际土壤及还田秸秆中Cry1Ab蛋白的田间残留降解动态,并分别用移动对数模型、指数模型和双指数模型对秸秆分解释放Cry1Ab蛋白的田间降解动态进行拟合,估算了DT5o和DT90值.结果表明:Bt玉米不同生育期根际土壤中Cry1 Ab蛋白含量差异较大,但总体随生育期的延长而显著降低.收获后地表覆盖和埋入土壤两种秸秆还田方式下,秸秆中Cry1Ab蛋白在土壤中的降解规律基本一致,均呈现前期大量快速降解,中后期极少量稳定降解两个阶段.秸秆还田7d内,地表覆盖处理的Cry1Ab蛋白降解率均极显著高于埋入土壤处理;10 d时两处理的降解率基本一致,分别为88.8％和88.6％;20 d后,两处理秸秆中Cry1Ab蛋白的降解日趋缓慢;至180 d时仍能检测到少量的Cry1Ab蛋白.3种模型均能较好地反映秸秆中Cry1Ab蛋白的田间降解规律,从相关系数(R)及DTgo值与实测值的吻合程度来看,双指数模型最优.     

转Bt基因玉米根际土壤及秸秆残体中

采用ELISA法研究了田间种植条件下转Bt基因玉米MON810生育期根际土壤及还田秸秆中Cry1Ab蛋白的田间残留降解动态,并分别用移动对数模型、指数模型和双指数模型对秸秆分解释放Cry1Ab蛋白的田间降解动态进行拟合,估算了DT₅₀和DT₉₀值. 结果表明: Bt玉米不同生育期根际土壤中Cry1Ab蛋白含量差异较大,但总体随生育期的延长而显著降低. 收获后地表覆盖和埋入土壤两种秸秆还田方式下,秸秆中Cry1Ab蛋白在土壤中的降解规律基本一致,均呈现前期大量快速降解,中后期极少量稳定降解两个阶段. 秸秆还田7 d内,地表覆盖处理的Cry1Ab蛋白降解率均极显著高于埋入土壤处理;10 d时两处理的降解率基本一致,分别为88.8%和88.6%;20 d后,两处理秸秆中Cry1Ab蛋白的降解日趋缓慢;至180 d时仍能检测到少量的Cry1Ab蛋白. 3种模型均能较好地反映秸秆中Cry1Ab蛋白的田间降解规律,从相关系数(R)及DT₉₀值与实测值的吻合程度来看,双指数模型最优.     

一株玉米秸秆纤维素降解放线菌的筛选、鉴定及酶学特性研究

为解决玉米秸秆固废污染和秸秆资源有效利用问题，采用刚果红染色法（水解圈法）和3，5-二硝基水杨酸（DNS）法从玉米秸秆还田土壤中筛选到一株纤维素降解菌，并对该微生物进行生理生化和分子生物学鉴定，发现该菌株降解纤维素效果较好，经鉴定该菌株为纤维素链霉菌（Streptomyces cellulosae），命名为SJS-15，并对该菌株的酶学特性及纤维素降解能力进行了初步研究。结果表明，菌株SJS-15在发酵培养基中的纤维素酶活（CMC）峰值为30.5 U/mL，最适反应pH为6.0，滤纸酶活（FPA）峰值为25 U/mL，最适反应pH为8.0，两种酶均能在温度20~60 ℃，pH 4.0~10.0范围内保持较高酶活性。纤维素分解实验表明菌株SJS-15对玉米秸秆和滤纸有分解能力，40 d时对玉米秸秆降解率为35.6%（质量分数，下同），对滤纸降解率为18.6%。扫描电镜结果显示经菌株处理的玉米秸秆较对照有明显降解痕迹。菌株SJS-15具有良好的抗逆性和玉米秸秆纤维素分解能力，可作为玉米秸秆还田和堆肥发酵的高效菌株进行进一步研究。     

三株高效秸秆纤维素降解真菌的筛选及其降解效果

【目的】利用多种筛选方法,获得高效秸秆纤维素降解真菌,并研究其秸秆纤维素的降解能力。【方法】采用滤纸片孔洞法、滤纸条降解法、羧甲基纤维素钠(CMC-Na)水解圈测定法、秸秆失重法、纤维素分解率测定法、胞外酶活测定法等常规秸秆纤维素降解菌的筛选方法。【结果】筛选到3株具有较强纤维素降解能力的真菌菌株,经初步鉴定菌株98MJ为草酸青霉(Penicillium oxalicum)、菌株W3为木霉(Trichoderma sp.)、菌株W4为扩张青霉(Penicillium expansum)。菌株W4具有非常强的秸秆纤维素降解能力,10d内对秸秆的降解率可达56.3%,对纤维素、半纤维素和木质素的分解率分别为59.06%、78.75%和33.79%。菌株W4的胞外纤维素酶活力在14.25-49.75U/mL之间。【结论】筛选获得3株高效秸秆纤维素降解真菌菌株,其中菌株W4的纤维素酶活高于已报道的菌株,是一株十分具有研究开发潜力的纤维素酶生产菌株。     

产纤维素酶耐高温真菌在有机肥料生产中的潜在价值

农作物秸秆是农作物生产中一种富含氮、磷、钾、钙、镁和有机质等有效成分的可再生资源,我国每年的农作物秸秆废弃物产量巨大,因此,如何有效地"转废为宝",实现对农作物秸秆废弃物的再利用一直是我国学者关注的问题.秸秆堆腐还田技术是有机废弃物再利用的理想途径之一,其作用原理是利用微生物的分解作用,促使农作物秸秆发酵腐熟后成为优良的绿色有机肥.农作物秸秆属于高纤维素含量废弃物,且纤维素的结构复杂、降解困难,如何加速纤维素的分解是实现农作物秸秆堆肥物料快速分解、达到腐熟的关键问题.已有研究表明,在堆肥中接种高温或耐高温降解菌可促进有机物降解,提高堆肥高温期温度,延长高温期,加快堆肥腐熟.     

两株戴氏霉对水稻秸秆的降解及产酶研究

本文旨在构建能够高效降解水稻秸秆的戴氏霉组合菌。通过兼容性试验,选取木质素降解菌灰戴氏霉H57.1菌株与纤维素降解菌合川戴氏霉H08.1菌株组合发酵降解水稻秸秆,以失重法和范氏洗涤剂法检测其对秸秆的降解效果,用胞外酶活测定法探索其产酶规律。结果表明,菌株H57.1和H08.1具有良好的兼容性;组合菌H57.1+H08.1对水稻秸秆的降解能力明显高于单一菌株,秸秆失重率高达55.7%,木质素、半纤维素和纤维素降解率分别为48.9%、72.6%和57.0%;对发酵过程产酶情况的分析进一步表明,组合菌H57.1+H08.1降解水稻秸秆的能力与其产酶能力密切相关。     

微生物菌剂对秸秆降解及秸秆中微生物变化规律的影响

为明确不同微生物菌剂对秸秆降解率及秸秆降解过程中秸秆周围微生物变化规律的影响，在温室大棚内进行不同微生物菌剂降解秸秆试验。以玉米秸秆为基质，设玉米秸秆（对照组）、玉米秸秆+哈茨木霉（Trichoderma harzianum）（处理1）、玉米秸秆+哈茨木霉+地衣芽胞杆菌(Bacillus licheniformis)（处理2）三个处理。研究结果显示，单独用哈茨木霉处理可显著提高秸秆降解率，前期作用尤其明显；在秸秆降解的前30 d，秸秆降解率与秸秆中可培养真菌、细菌、放线菌呈显著正相关。不同微生物菌剂对秸秆中可培养微生物数量变化有显著影响，单独接种哈茨木霉后，秸秆中可培养真菌、细菌、放线菌数在前30 d显著高于对照组和混合菌剂处理，不同处理可培养活菌数均在90～120 d达到峰值，然后开始下降。研究结果表明，不同微生物菌剂对秸秆降解有显著影响，单独用哈茨木霉处理可显著提高秸秆降解率；不同处理对秸秆中可培养真菌、细菌、放线菌的影响有显著差异。可为秸秆降解与微生物相关性研究提供参考。     

降解菌系和助腐剂对不同还田方式下水稻秸秆降解特征的影响

基于田间小区试验,设置两种秸秆还田处理(以秸秆混土处理模拟旋耕还田,以秸秆不混土处理模拟秸秆沟埋还田),采用尼龙网袋法,通过测定秸秆降解率及秸秆半纤维素、纤维素和木质素的含量,研究两种还田方式下施用降解菌系和助腐剂后水稻秸秆的降解规律。结果表明:在试验前5个月,不混土处理的秸秆降解率始终高于混土处理4.5%～11.6%;至第12个月,混土处理秸秆降解率显著高于不混土处理4.9%～13.0%。在前2个月,各处理水稻秸秆降解较快,秸秆降解率均超过50%,此后秸秆降解进入缓慢期。混土处理中,降解菌系和助腐剂仅能在第1个月提高秸秆的降解效率,此时秸秆降解率分别比对照组高11.0%和10.2%(P0.01);不混土处理中,降解菌系在1～5个月内显著提高秸秆降解效率4.3%～9.7%,助腐剂在1～12个月显著提高秸秆降解效率7.3%～14.4%。试验至第5个月时,喷施助腐剂的秸秆半纤维素、纤维素和木质素的降解率分别达79.1%、80.2%和34.7%,且与喷施菌系处理差异不显著。综上所述,秸秆降解菌系和秸秆助腐剂均能提高水稻秸秆的降解效率,且不混土处理的施用效果好于混土处理。     

微生物菌群协同提高水稻秸秆转化机制的解析

单一微生物降解水稻秸秆效果不明显,多种微生物组合在一起的微生物菌群能够有效降解水稻秸秆,是当前降解秸秆类废物的首要选择。【目的】探究微生物菌群比单一菌株转化秸秆效率提高的种间协作机制,为改善秸秆类物质的生物降解过程提供理论参考。【方法】通过菌种组合,以水稻秸秆减重率为指标人工构建出降解效果优于单菌培养的微生物群体组合,利用分子生物学方法对实验菌株进行鉴定,结合纤维素酶活性、发酵产物GC-MS分析等指标得出水稻秸秆降解时微生物群体的种间协作机制。【结果】菌株B (Bacillus cereus)在30℃培养8 d水稻秸秆降解率为50.9%,菌株B与菌株D_2(Bacillus amyloliquefaciens)、W_1(Ochrobactrum intermedium)、G_1降解(Bacillus licheniformis)组合构成4株菌BD_2W_1G_1的复合菌群,在30℃培养8 d水稻秸秆降解率为73.3%,比B单菌株分解能力提高22.4%。发酵产物分析结果表明,菌株B单独培养产生大量酸、酚等物质。B+D_2联合培养,发酵液内酸类相对减少87.4%,酚类相对减少61.9%(十五烷酸、正十六烷酸、2,4二叔丁基酚和6-叔丁基对甲酚减少明显),水稻秸秆降解率为64.6%。当B+D_2+W_1联合培养,发酵液内酚类进一步减少15.7%(6-叔丁基对甲酚减少明显),水稻秸秆降解率为71.0%。当B+D_2+W_1+G_1联合培养,酚类继续减少10.7%,水稻秸秆降解率为73.3%。【结论】十五烷酸、正十六烷酸、2,4二叔丁基酚和6-叔丁基对甲酚等酸、酚类物质会抑制菌株B分解水稻秸秆的效率,菌株D_2、W_1、G_1能够减少包括4种物质在内的酸、酚类含量(酸类共减少82.9%,酚类共减少88.2%),来提高分解效率(共提升22.4%的分解效果)。水稻秸秆成分复杂,具有不同功能的菌种组合成复合菌剂,减少反馈抑制形成代谢互补,能有效提高水稻秸秆生物降解效果。     

Bt玉米秸秆Bt蛋白的土壤降解及其拟合模型的比较

研究4种Bt玉米(34B24、NK58-D1、R×601RR/YG和农大61)秸秆分解释放Bt蛋白的土壤降解动态,分别应用一级动力学反应方程、双指数模型和移动对数模型进行了拟合。结果表明,4种Bt玉米Bt蛋白的土壤降解均呈现前期负指数大量快速降解和中后期极少量稳定两个阶段,双指数模型与移动对数模型的拟合结果比一级动力学反应方程更符合实际。判断模型拟合精度时不能只用拟合结果的统计指标值来确定,图示对比实测值与模拟值的差异是避免应用统计学拟合精度指标很高但并不符合实际的模型错误估算DT50的有效办法。在研究转基因作物秸秆中Bt蛋白土壤降解规律时,应该根据其土壤降解前期较快的特点,增加前期取样的次数,以便更好地对模拟模型求得的安全性评价关键参数DT50的准确性进行科学判断。     

Homogenous reactions of cellulose from different natural sources

Two different homogenous reactions on bacterial cellulose (BC), kenaf fiber (KF) and microcrystalline cellulose (MC) were performed to monitor their chemical reactivity. The first reaction was selective oxidation of the primary hydroxyl group with sodium chlorite in the presence of catalytic amount of sodium chloride. While, the second was the formation of triester hypoiodous cellulose using potassium iodate and potassium iodide. The chemical structures of these derivatives were investigated using FT-IR and solid state ¹³C NMR spectroscopies. The BC fibrils required the shortest time among these cellulose samples for both reactions, whereas the viscosity values of BC after iodination and oxidation have the best values compared to KF and MC. FT-IR results show the absence of the hydroxy group of BC and a weak absorption band in both KF and MC. On the other hand, the crystallinity index (CI) of BC is higher than those of both KF and MC. FT-IR spectra of the oxidized different cellulose samples, confirmed the presence of a strong absorption band at around 1590 cm⁻¹ that attributed to vibration band of carbonyl group of carboxylic moiety. Moreover, in the ¹³C NMR spectrum of oxidized cellulose, the lack of signal at 62 ppm and the appearance of signal at 171 ppm indicated that the primary alcohol group is completely oxidized to carboxylic acid. These results showed that BC had a higher reactivity than other samples due to its great purity and low degree of polymerization.     

纤维素分解菌对不同纤维素类物质的分解作用

经过CMC平板、滤纸液化和摇瓶培养试验 ,发现 6株菌中 ,产黄纤维单胞菌 (CellulomonasFlav igena)和康氏木霉 (Trichodermakonigii)分解纤维素类物质的能力比较强 ,对来源不同的纤维素类物质分解能力差异很大 ;真菌与细菌一起接种时 ,分解纤维素类物质的速度明显高于其中任何一个单一菌株 ,说明纤维素类物质的分解需要多种微生物的联合作用     

RNA sequence determination in the nanogram range by a combination of in vitro labeling procedures.

Recently developed methods which allow one to read RNA sequences directly from polyacrylamide gels do not always provide unequivocal results. A combination of primary and secondary in vitro 5′-labeling, as presented here, is methodically and in its results equivalent to fingerprinting and sequencing techniques developed for in vivo labeled RNA. 5 S RNA was used to demonstrate the applicability and reliability of this combination of postlabeling procedures: 5 μg RNA was partially digested, and the resulting overlapping fragments were 5′-³²P-labeled with T4 phage-induced polynucleotide kinase in vitro. After two-dimensional polyacrylamide gel electrophoresis and carrier-free electrophoretic elution, the labeled long fragments, obtained in the 10-ng range, were completely degraded with RNase T₁ and RNase A, respectively. These digests were again ³²P-phosphorylated with T4 kinase and lead to fingerprints which allowed the deduction of the nucleotide sequences of the corresponding long fragments.     

The three-dimensional morphology of aggregates of native cotton cellulose microfibrils

The three-dimensional morphology of native bacterial cellulose is confirmed by scanning electron microscopy. In addition, it is shown by scanning electron microscopy, and transmission electron microscopy with positive staining by phosphotungstic acid ions that aggregates of microfibrils of native cotton cellulose have a similar structure. The results are consistent with previous reports on microfibrils of algal cellulose. These observations exclude a simple spinneret process as a mechanism of formation of the microfibrils of these sources of cellulose.     

植物纤维素生物合成及其相关酶类

纤维素是由成千上万个D-葡萄糖分子通过β-1,4糖苷键连接的具有一定立体构象的链状聚合物,其葡萄糖残基约为2000～25000个。它是细胞壁的主要组成成分之一。植物,大多数藻类,一些细菌和真菌甚至有些动物都能合成纤维素。它作为世界上最丰富的,具有巨大商业价值的生物多聚体,几十年来一直受到人们的重视,成为人们的研究热点。尽管如此,这方面的研究仍比较滞后,人们对纤维素生物合成途径及其相关酶类还是知之甚少。近几年来,随着基因组学的发展,关于纤维素的生物合成及相关基因表达调控的研究也成绩斐然,现就高等植物纤维素生物合成途径及其相关的纤维素合成酶的最新研究进展作一介绍。     

Modification of carboxymethyl cellulose through oxidation

The periodate oxidation reaction of carboxymethyl cellulose involve the primary and secondary hydroxyl groups of the pyranose ring. The reaction is accompanied by the opening of the pyranose ring and resulting product is dialdehyde carboxymethyl cellulose along with some hydrated forms. In this process the glucosidic bond becomes weaker; the formation of carboxyl groups induces a depolymerization, thus reducing the polymerization degree and the physical and mechanical strength of the material. The reaction has been has been carried out at pH 3.5, temperature 45 °C for 0.5-4 h.     

裂褶菌纤维二糖脱氢酶吸附纤维素性质

纤维二糖脱氢酶（ＣＤＨ）可以吸附棉花、微晶纤维素和酸处理纤维素,４ｍｉｎ便都达到平衡。与纤维素酶明显不同,该酶的Ｓｃａｔｃｈａｒｄ吸附曲线都是一条直线,为典型的单结合位点模型（ｏｎｅ－ｂｉｎｄｉｎｇ－ｓｉｔｅ ｍｏｄｅｌ）。观察到ｐＨ值、温度、乙二醇和ＮａＣｌ对ＣＤＨ吸附微昌纤维素有影响,并进行了讨论。     

Compression, compaction, and disintegration properties of low crystallinity celluloses produced using different agitation rates during their regeneration from phosphoric acid solutions

The tabletting characteristics of low crystallinity celluloses (LCPC)-LCPC-700, LCPC-2000, and LCPC-4000-prepared using agitation rates of 700, 2000, and 4000 rpm, respectively, during their regeneration from phosphoric acid, were evaluated and compared with those of Avicel PH-102 and Avicel PH-302. The mean deformation pressure values calculated from the linear region of the Athy-Heckel curves indicated LCPC-4000 to be the most ductile material. The area under the Athy-Heckel curve for LCPC-4000 was 330 MPa, whereas LCPC-700 and LCPC-2000 showed a corresponding value similar to that of Avicel PH-102 and Avicel PH-302 (192–232 MPa). The tensile strength of LCPC and Avicel compacts increased linearly with increasing applied pressures. A comparison of the area under the tensile strength-compression pressure curves indicated that LCPC-4000 formed the strongest tablets. The strengths of LCPC-700 and LCPC-2000 compacts, in contrast, were slightly lower than that of Avicel PH-302 and Avicel PH-102, respectively. The compacts of both LCPC-4000 and Avicel PH-102 were intact in water for 6 hours, whereas LCPC-2000 and Avicel PH-302 compacts disintegrated in 4 minutes and 2 minutes, respectively. In conclusion, LCPC-4000 was the most ductile material and exhibited the highest compression and compaction characteristics. The corresponding properties of LCPC-700 and LCPC-2000, in contrast, were comparable to that of Avicel PH-102 or Avicel PH-302.     

Disappearance of the Cuticle and Wax in Outermost Layer of Callus Cultures and Decrease of Protective Ability against Microorganisms

In electron microscopic observation, neither wax nor cuticle was observed on the outermost layers of callus tissues. Chemical estimation of wax in the callus surface was attempted by thin-layer chromatography of solvent extracts of callus tissues in comparison with those of barley and rice leaves. Hydrocarbons and free alcohols were detected in lyophilized callus tissues, but no wax esters or ketones were detected. Germination test indicated that germination of spores of Aspergillus oryzae was less favored on hydrophobic membranes than that of spores of Alternaria sp. and Botrytis cinerea.

From these results, we inferred that the lack of cuticle and wax in the outermost layer of callus tissues facilitated spore germination and penetration, and A. oryzae, a saprophytic fungus, could also readily penetrate into callus tissues.