木霉菌(Trichoderma spp.)嗜铁素具有吸收、贮存和胞内运输铁的功能,对菌体自身的生长发育有重要作用,也是重要的生防促生因子,但嗜铁素的产量会受到环境中多种胁迫因子的影响.真菌的钙调磷酸酶基因能够调控外源离子的胁迫反应,但钙调磷酸酶信号通路与嗜铁素合成之间的关系并不明确.以棘孢木霉菌(Trichoderma asperellum)T6为材料,研究了NaCl、CaCl2及pH对棘孢木霉菌T6嗜铁素产量以及嗜铁素合成关键基因(SidA)与钙调磷酸酶催化亚基基因(CnA)表达量的影响.结果 表明,高浓度NaCl和CaCl2以及高pH均能抑制嗜铁素的产生.利用半定量PCR分析得知,CnA基因与SidA基因之间存在一定的负调控关系,该结果与钙调磷酸酶专性抑制剂环孢菌素A能够提高嗜铁素产量的结果相一致.

2003年, 我博士毕业来到山东师范大学, 承担了两个专业两门专业课的主讲任务, 其中一门是生物技术专业大三学生每年春天上的"酶工程". 经过六年的教学积累,2009年寒假, 我不声不响地酝酿并完成了一项在旁人看来不可思议、但我却认为是一项使命的工作——酶工程专业课的双语教学,从此,有了我与众多生技学子之间亦师亦友、教学相长的故事,尤其是那绚丽的"五朵金花"常浮现在我的眼前.

高等植物细胞内囊泡运输介导了膜结构之间的物质运输和信号转导,其中,胞吐调控了运输囊泡向质膜的转运,参与了植物细胞壁形成、细胞分泌和环境响应等多种生物学过程.胞吐可以通过重构及强化细胞壁阻止病原体定殖、分泌抗微生物因子抵御病原体的入侵以及调控植物激素载体蛋白和受体蛋白的极性运输等参与抗病应答.遗传学证据表明胞吐是调控植物生物胁迫响应的重要机制.该文综述胞吐参与植物生物胁迫应答分子机制的研究进展,以期为本领域研究提供参考.

In order to study the effects of chitoson-alginate composite membrane on cotton seed germination and growth and development of seedlings,the cotton seeds were conducted coating treatment with composite membrane of concentration ratios of chitoson to alginate as 5∶ 1,4∶ 1,3∶ 1,2∶ 1,1∶ 1,0∶1 respectively.The results showed that,when the concentration ratio reached 3∶1 and 4∶ 1,the properties of composite membrane were the best.The seed germination rate,contents of chlorophyll,soluble sugar and soluble protein of cotton seedlings were all more than those of no coating treatment and single reagent treatment.They firstly increased and then decreased with the decrease of concentration ratio,and the effect was the best when the concentration ratio was 3∶ 1.

Stimulating students'interest in learning and activating classroom atmosphere as well as establishing an efficient teaching model have long been the important contents of higher education reform. During the past 7 years, bilingual lecturing of all-English courseware and other interesting methods have been adopted in undergraduate courses of "Enzyme Engineering" and "Fermentation Technology" in Shandong Normal University. These methods have been found to be effective to stimulate the students'learning enthusi-asm and to help them gain knowledge and skills. The innovative teaching methods have been well received by the students and will be helpful in biological education reform in universities.

A strain of Trichoderma spp.LT19 was isolated from soil in Shandong Province,which showed a broad inhibition spectrum against various soil-borne plant phytopathogenic fungi.It also exhibited some plant growth-promoting attributes such as phosphate solubilization,indol-3-acetic(IAA) productivity,1-aminocyclopropane-1-carboxylate(ACC) deaminase activity and siderophore(S) synthesis ability.The pot experiments showed that inoculation with the strain could increase the biomass of cucumber seedlings.Based on its morphology and ITS gene sequences,the strain LT19 was identified as Trichoderma asperellum.

转基因玉米MIR162是美国先正达公司开发的新型抗鳞翅目昆虫的转基因玉米品种.本研究目的在于建立该品系转化事件特异性定性、定量检测方法并形成定性检测国家标准.定性PCR体系经实验室内部比对及实验室间循环验证结果表明,该体系特异性强,灵敏度较高(0.1％),具有良好的重复性和再现性.TaqMan探针实时荧光PCR检测结果表明,所设计的探针特异性强,检测低限(LOD)在0.01％以下;定量标准曲线R2均为0.999,扩增效率为98.622％和99.602％,SD范围为0.014～0.209,RSD的范围为0.049％～0.879％,稳定性好;对转基因含量为1.0％、0.5％、0.1％的样品定量结果相对偏差在8.0％以内,结果较为准确.本研究建立的方法完全可用于含转基因玉米MIR 162成分样品的检测.

采用SYBR Green实时荧光定量PCR技术,建立转基因大豆BPS-CV127-9的定量检测方法.通过设计特异引物,扩增内标准基因lectin和BPS-CV127-9的5’侧翼序列,建立2种基因的拷贝数-CT标准曲线,根据标准曲线方程计算样品中的转基因含量,并且通过熔解曲线分析扩增反应特异性.结果表明,lectin基因和侧翼序列标准曲线线性关系良好,R2值分别为0.999和0.998,变异系数(CV) 1.50％～18.51％、标准偏差(SD)0.02 ～0.07.检测4个已知BPS-CV127-9含量(1％、0.5％、0.1％、0.05％)的转基因混合样品,实测值与实际值接近.该检测方法具有快速、灵敏、准确、特异、高通量等优点,可以作为转基因大豆BPS-CV127-9的定量检测方法.

In order to evaluate the effect of transgenic maize on epiphytes, the transgenic Bt maize Mon810 and its parental non-transgenic maize were grown in field to study the quantitative changes of culturable microorganisms and the diversity of bacterial functional groups at different growth stages in 2010 and 2011.Although there were differences in the colony-forming unites of culturable bacteria, actinomycetes and fungi of phyllosphere between Bt maize and non-Bt maize in different growing periods and different years, the trend of annual differences was basically consistent at the same growth period: the quantity of microbes in seedling stage is fewest and reaches the peak in the late of growing period.Compared with the controls, transgenic Bt maize seemed to stimulate the reproduction of phyllosphere fungi.However, no significant effect on the populations of actinomycete was observed.Significant differences were detected for bacteria at jointing stage, trumpet stage, silking stage and milking stage.In 2010, the Shannon-Wiener index, Simpson index and evenness index of bacteria physiological groups were higher at seedling, jointing and full-ripe stage, but lower at trumpet stage, tasseling stage, silking stage and milk stage.In 2011, the population characteristic parameters of the microorganisms of Bt-corn are more than the ones of non-Bt corn in the whole growing period except trumpet stage.It may be concluded there was some difference in terms of impacts between transgenic maize and non-transgenic maize, while the main impacts on microbial community composition were likely due to different years and different growth periods.

With the transgenic insect-resistant corn Mon810 and its non-transgenic parent as materials,the enzyme-linked immunosorbent assay(ELISA) method was used to detect the expression and quantitate the exudation of Bt protein in corn leaves.The results showed that Bt protein could be detected in tissue and on surface of leaves.The Bt content in tissues showed a rising trend from seedling stage to silking stage with the advances of vegetable growth,and reached the peak at the silking stage and then declined gradually.The variation of Bt exudation on leaf surface had no obvious regularity.

One strain of biocontrol Trichoderma spp.Q3 with the function of pesticide degradation was screened from 9 antagonistic Trichoderma spp.strains preserved in our laboratory.It could utilize 100 mg/L beta-cypermethrin,chlorpyrifos and imidacloprid as the sole carbon source for growth and the degradation rate reached 90.43%,86.71% and 78.81% respectively within 7 days.The residual dynamics and final residues of beta-cypermethrin and chlorpyrifos in Brassica campestris L.were studied in greenhouse and determined by gas chromatography.The results showed that Trichoderma spp.Q3 could significantly decrease the residue of the two pesticides in Brassica campestris,and the degradation rate of beta-cypermethrin and chlorpyrifos was 45.54% and 72.58% respectively after 17 days.

Based on the specific structures of tomato endogenous reference gene PG and general genes of transgenic tomato,such as CaMV 35S promoter,NOS terminator,npt-Ⅱ gene and antisense EFE gene,five pairs of primers were designed for qualitative PCR.The results showed that the target DNA sequences specific to every transgenic tomato could be detected,which indicated that the primers had high specificity and accuracy,and could identify transgenic tomatoes specifically.

With the increase of planting area of transgenic Bt crops year by year,people paid more and more attention to the potential safety of transgenic crops release.Bt protein could be released into soil by transgenic Bt crops through plant residues and root exudates,and its enrichment could affect the enzyme activities and fertility of soil.But the influence was uncertain with different Bt crops and soil conditions.Future observation and research should be conducted in a long term on the effects of commercialized Bt crops on soil enzyme activities and soil fertility.In this paper,the advances in research on development of transgenic Bt crops and the influences of their cultivation and straw decomposition on soil enzyme activities and fertility were reviewed to provide reference for ecological assessment of transgenic Bt crops to soil environment.

Transgenic event-specific primers and Taqman probes based on the left-flanking sequence of MON88017 and endogenous gene (zSSIIb) were designed, and a real-time PCR method was developed to quantitatively detect the event-specific genetically modified maize. The reference molecule composed of endogenous gene and flanking sequence of MON88017 was constructed artificially. Two standard curves of the reference gene sequence and the 5' flanking sequence were established. We detected five mixed samples, and their genetically modified contents of MON88017 were 0.01%, 0.05%, 0.1%, 0.5%, and 1%, respectively. The lowest detection limit was 19-30 copies. This study indicated that the Taqman probes real-time PCR is highly specific and sensitive.

魏晋异代之际,社会动乱异常,不少文人志士皆丧命于政治漩涡之中,阮籍,中国文学史上的一朵奇葩,以其独特的中庸处世之道不仅保全性命并使自己的人生价值得到实现.其秉承老、庄,但又独具特色的中庸思想在其政治、生活和文学创作中可见一斑.

The experiment was conducted to investigate the integration site of transgene in maize MON88017 and to establish event specific methods for qualitative detection of MON88017 based on the left border junction fragment, which was isolated with the amended GenomeWalker and Nested-PCR methods. Sequence alignment between the T-DNA sequence and isolated junction fragments showed a 504 bp junction fragment of MON88017 including 336 bp of T-DNA sequence and 168 bp of MON88017 genome DNA. MON88017 event-specific qualitative PCR method was established with the primers (MON88017-1F/R) targeting the junction regions to produce a 446 bp product. The limit of detection for qualitative PCR assay was 0.1%. The method developed in this work is highly specific, sensitive and suitable for MON88017 sample detection.

Trichoderma spp.are effective biocontrol agents for plant pathogens,and some strains are also known for their abilities to colonize plant roots to cause substantial changes in plant metabolism.This review summarizes recent advances in research of Trichoderma spp.root colonization,which promote plant growth by producing plant growth regulators,inhibiting or reducing detrimental substances in rhizosphere and increasing plant nutrient intake.They also produce elicitors to induce localized or systemic resistance responses by inducing plants to form wall appositions and synthesize antimicrobial compounds.This overview serves as a reference to extend the application of Trichoderma spp.in agricultural production.

建构主义认为,知识是学生在特定的情境之下,借助于教师的帮助和学习材料,通过意义建构而获得的.学生能够亲身体验学习内容,才能提高学生的积极性,发挥学生的创造性.知识的学习由被动接受转变成主动探索的过程,学习的效果大为提高.

One dominant endophytic bacterial strain W7 was isolated from the root of fresh leek,which had a broad inhibition spectrum against several vegetable pathogenic fungi.The results of its antagonistic mechanism showed that the extracellular metabolites had no effect on pathogenic fungi,but the somatic polysaccharose which obtained from cells by ultrasonic disruption and organic solvent precipitation could inhibit the growth of mycelium.Strain W7 was also found to be capable of utilizing beta-cypermethrin as the sole source of carbon for growth,and its degradation within 7 days at initial concentration of 100 mg/L was 51.3%.The results indicated that strain W7 was a pesticide-degrading and an endophytic bacteria suitable for biocontrol.It was identified as Paenibacillus spp.based on morphological,physiological-biochemical properties and 16S rDNA sequence analysis.

In this study,yeast invertase(β-fructofuranoside hydrolase) was extracted and purified by cell disruption,heat denaturation,organic solvent precipitation and DEAE-Sepharose Fast Flow chromatography.Five chemical modification agents(PMSF,DTT,SUAN,NBS,EDTA) and several other metal ions were used to determine the amino acid residues involved in the active site of yeast invertase.The result indicated that serine residues and metal ions were inessential to the invertase activity while the lysine and cysteine residues were linked to the enzyme activity, which were not in the active site of the enzyme.That the indolyl ring of tryptophan residues could be modified by N-bromosuccinimide and these groups could be protected by sucrose revealed that the tryptophan residues were present at the invertase active site.