Xα21基因导入水稻及转基因植株的鉴定

以水稻愈伤组织为受体、质粒pCXK1301为Xα21基因供体，用农杆菌EHA105和PDS－1000／He基因枪转化受体，经Hyg浓度梯度筛选，获得一些转基因水稻植株，GUS检测、PCR扩增和Southern杂交分析表明，报告基因（GUS）、选择标记基因（Hyg^r）和抗白叶枯病基因Xα21都已整合入转化水稻基因组中，Southern杂分析显示了Xα21基因在转化水稻基因组中整合的拷贝数。田间接菌鉴定表现抗白叶枯病。     

Use of reporter-gene based bacteria to quantify phenanthrene biodegradation and toxicity in soil

A phenanthrene-degrading bacterium, Sphingomonas paucimobilis EPA505 was used to construct two fluorescence-based reporter strains. Strain D harboring gfp gene was constructed to generate green fluorescence when the strain started to biodegrade phenanthrene. Strain S possessing gef gene was designed to die once phenanthrene biodegradation was initiated and thus to lose green fluorescence when visualized by a live/dead cell staining. Confocal laser scanning microscopic observation followed by image analysis demonstrates that the fluorescence intensity generated by strain D increased and the intensity by strain S decreased linearly at the phenanthrene concentration of up to 200 mg/L. Such quantitative increase and decrease of fluorescence intensity in strain D (i.e., from 1 to 11.90 ± 0.72) and strain S (from 1 to 0.40 ± 0.07) were also evident in the presence of Ottawa sand spiked with the phenanthrene up to 1000 mg/kg. The potential use of the reporter strains in quantitatively determining biodegradable or toxic phenanthrene was discussed.     

用12SrRNA基因序列研究潮龟科（BATAGURIDAE）闭壳龟类的进化

测定了潮龟科（Ｂａｔａｇｕｒｉｄａｅ）具闭壳结构的齿缘摄龟属（Ｃｙｃｌｅｍｙｓ）、锯缘摄龟属（Ｐｙｘｉｄｅａ）、闭壳龟属（Ｃｕｏｒａ）共６种线粒体１２ＳｒＲＮＡ基因片段的序列，结合潮龟科其它一些属的序列数据，探讨了具闭壳结构龟类的起源与进化以及闭壳龟属的分类学问题．认为闭壳龟属与锯缘摄龟属为姐妹群，两者与齿缘摄龟属的亲缘关系较远，提示这些具闭壳结构的龟类可能不是由一个共同祖先分化而来．本文的结果不支持将闭壳龟属拆分为闭壳龟属和盒龟属两属     

家蚕斑纹限性品种SC_1的RAPD差异DNA片段的克隆及其酶切图谱分析

利用RAPD技术对家蚕限性普斑品种SC1的雌雄体分别进行PCR扩增通过120种引物的筛选，得到1个雌体特异的RAPDDNA片段将该片段克隆到pUC19质粒，井进行了限制性内切酶图谱分析.     

Monitoring aromatic hydrocarbon biodegradation by functional marker genes

The development of biological treatment technologies for contaminated environments requires tools for obtaining direct information about the biodegradation of specific contaminants. The potential of functional gene array analysis to monitor changes in the amount of functional marker genes as indicators of contaminant biodegradation was investigated. A prototype functional gene array was developed for targeting key functions in the biodegradation of naphthalene, toluene and xylenes. Internal standard probe based normalization was introduced to facilitate comparison across multiple samples. Coupled with one-colour hybridization, the signal normalization improved the consistency among replicate hybridizations resulting in better discrimination for the differences in the amount of target DNA. During the naphthalene biodegradation in a PAH-contaminated soil slurry microcosm, the normalized hybridization signals in naphthalene catabolic gene probes were in good agreement with the amount of naphthalene-degradation genes and the production of 14CO2. Gene arrays provide efficient means for monitoring of contaminant biodegradation in the environment.     

连续施用发酵猪粪对土壤中四环素抗性基因数量的影响

为了研究连续施用发酵猪粪之后,土壤中抗性基因的数量以及发酵猪粪的施用量对土壤抗性基因的影响,采用实时荧光定量PCR方法,对稻麦轮作模式下连续6年施用4.5t/hm2和9.0t/hm2发酵猪粪的土壤进行了四环素抗性基因(TRGs)的检测和定量分析.在施用发酵猪粪的土壤中能够检测到9种TRGs.施用发酵猪粪显著增加了土壤中tet G、tet L、tet B(P)、tet O、tet W的绝对数量,其中tet B(P)、tet W、tet O的绝对数量受到了施肥用量的影响;而tet Z、tet C和tet S的绝对数量则没有受到施肥的影响.上述8种TRGs在同一施肥处理的0~5cm、5~10cm以及10~20cm土壤中的相对丰度分布均无显著差异.发酵猪粪的施用也显著增加了tet G、tet L、tet B(P)和tet O的相对丰度,但是仅有tet O的相对丰度受到了施肥用量的显著影响.本研究表明,在稻麦轮作模式下,农田土壤中的TRGs数量会受到发酵猪粪中残留TRGs的影响,连续施用该粪肥会显著增加土壤中tet G、tet L、tet B(P)和tet O的绝对数量和相对丰度.因此,需要优化畜禽粪堆肥发酵工艺,以减少抗性基因在粪肥中的残留.     

微生物聚磷及其酶学调控

本文论述了微生物细胞内磷酸盐转运机制和与聚磷相关的酶—PPK、PPX及两者对微生物聚磷行为的调控,并对今后研究重点做出展望.论文认为对于微生物聚磷的研究应从纯种微生物转移到强化生物除磷系统的混合微生物菌群,运用分子生物学和生物信息学手段分析聚磷优势菌种的PPK和PPX酶的结构特征及其在活性污泥中的变化规律,进一步深入认识生物除磷调控机理,开辟寻求提高系统除磷效率的新途径.     

Interspecific and intraspecific identification of Dendrobium based on the psbAtrnH intergenic region sequences and the 5S rRNA gene spacer sequences北大核心CSCD

This research aimed to investigate the interspecific and intraspecific identification of Dendrobium by using the multi-locus method so as to provide a molecular basis for Dendrobium identification through the combination of chloroplast psbA-trnH intergenic region sequences and ribosome 5S rRNA gene spacer sequences. PCR direct sequencing was applied to detect the chloroplast psbA-trnH intergenic region sequences as well as the ribosome 5S rRNA gene spacer sequences of 12 Dendrobium species, while the psbA-trnH intergenic region sequences of Dendrobium denneanum dq-2 variety and dq- 5line were cloned and sequenced for single nucleotide polymorphism (SNP) analyzing. The sequences were analyzed by the software Sequencher4.14, Bioedit7.0, MEGA5.2 and Dansp5.0; the interspecific and intraspecific Kimara-2-Parameter(K2P) distances were also calculated. The phylogenetic tree (using Neighbor joining method) was constructed with Bulbophyllum odoratissimum and Bletilla striata as outgroup. The results showed an average length of chloroplast psbA-trnH gene sequences in Dendrobium as 742.3 bp, with 72 variable sites, including 33 information sites; the average length of the ribosome 5S rRNA gene spacer sequences in Dendrobium was 336.4 bp, with 213 variable sites including 139 information sites. Using psbAtrnH intergenic region sequences in combination with ribosome 5S rRNA gene spacer sequences can not only identify D. denneanum, D. hancockil, D. thysiflorum, D. devonianum, D. moniliforme, D. chrysotoxum, D. officinale, D. heterocarpum and D. nobile, but also differentiate D. officinale from different geographical populations, and distinguish the dq-2 variety and dq 5line with SNP in the multi locus of D. denneanum.     

基于esp基因和JCV标记的MST方法在五大流域典型水源地中的应用

非点源污染目前已经成为影响水体环境的重要污染。微生物源示踪技术(microbial source tracking,MST)是解决非点源污染的一项新技术,它可以确定污染的宿主来源。肠球菌esp基因和多瘤病毒JCV可以作为检测水体中人源粪便污染的分子标记,其灵敏度和特异性都很高。为分析五大流域水源地是否受到人源粪便的污染,对辽河、海河、淮河、长江和黄河五大流域典型水源地水样进行采集和检测,选取人源粪便特异病原微生物肠球菌的esp基因和多瘤病毒JCV建立了相应的MST分子检测方法。结果表明,五大流域的典型水源地采样点均有可能受到了人源粪便的污染,可为当地相关部门提供技术支持和数据参考。