来自易变山羊草抗禾谷孢囊线虫基因的克隆与序列分析

禾谷孢囊线虫（Heterodera avenae）是严重危害禾谷类作物的病原线虫之一,它广泛分布于澳大利亚、欧洲、北美、印度和中国等世界主要小麦产区,使作物严重减产,造成巨大的经济损失。目前最有效的防治措施之一是将外源抗性基因导入栽培小麦(Triticum aestivum L.)，培育抗禾谷孢囊线虫的新品种。但迄今为止抗禾谷孢囊线虫基因克隆研究的相关报道却很少。 本实验根据此前从抗禾谷孢囊线虫材料E-10扩增得到的与来自节节麦（Aegilops tauschii）的抗禾谷孢囊线虫基因Cre3高度同源的序列Rccn4,设计出三条嵌套引物,采用SON-PCR(single oligonucleotide nested PCR)方法,从E-10基因组DNA中得到一个长为1264 bp的扩增产物(命名为Rccn-L),测序比对结果显示,这一序列将Rccn4的3’端延伸了1209 bp,与抗禾谷孢囊线虫Cre3基因核苷酸同源性为86﹪，核苷酸编码区长1026 bp，含一个不完整的开放阅读框,一个终止密码子，没有起始密码子和内含子结构,编码一个342个氨基酸残基的蛋白质。该蛋白质等电点为5.19，分子量为38112.6Da。从序列的第113位开始到第332位是NBS-LRR类抗病性基因LRR区,呈现XXLXXLXXL重复。LRR编码区内亮氨酸残基的含量达17﹪，与抗禾谷孢囊线虫Cre3基因LRR编码区的核苷酸和氨基酸同源性分别为89﹪和78﹪。本实验首次将SON-PCR成功地运用于植物基因克隆，为植物基因克隆提供了又一有效方法。 此外,还根据Cre3基因及其他的NBS-LRR类植物抗性基因的NBS和LRR区保守序列设计了两对特异性引物，从禾谷孢囊线虫抗性材料易变山羊草基因组DNA中扩增到两个相应的目标条带。测序分析结果表明，它们的长度分别为532bp和1175bp，构成了一个有32bp的共同序列的NBS-LRR编码区。其序列总长为1675bp（命名为RCCN），含有一个不完整的开放阅读框，没有起始密码子、终止密码子和内含子结构。其中编码序列为1673bp，可编码一个557个氨基酸的蛋白质，等电点（pI）为5.39，分子量为63537.5Da。与Cre3的核苷酸和氨基酸同源性分别为87.8﹪和77﹪。RCCN氨基酸序列中含有已知抗病基因NBS区域的几个保守模体：kinase2区的ILDD、kinase3的（ⅰ）ESKILVTTRSK，（ⅱ）KGSPLAARTVGG，（ⅲ）RRCFAYCS及EGF。RCCN NBS区与Cre3 NBS区的核苷酸和氨基酸的同源性分别为96.4﹪和94﹪。从氨基酸序列的274位到548位为LRR保守区，呈现不规则的aXXLXXLXXL（其中a代表I，V，L，F或M）重复，其中亮氨酸的含量为15.6﹪。该区域与Cre3的LRR区的核苷酸和氨基酸同源性分别为80.8﹪和74﹪。推测该序列可能为一个抗禾谷孢囊线虫的新基因。 本文对抗禾谷孢囊线虫基因的克隆研究，为进一步克隆基因全序列，探索其结构与功能，和研究该基因表达与调控提供了关键信息。同时也为通过基因工程途径将抗性基因向优良小麦品种高效、定向转移，最终培育出小麦抗禾谷孢囊线虫新品种奠定了基础。 Cereal cyst nematode (CCN) is a damaging pathogen of broad acre cereal crops in Australia, Europe, North America, India and China. It affects wheat, barley, oat and triticale and causes yield loss of up to 80%. At present, Transferring resistance genes against CCN into wheat cultivars and breeding varieties are considered one of the most effective methods for controlling the CCN. However, there are very limited reports concerning the cloning studies of resistance genes against the cereal cyst nematode. According to the sequence of Rccn4 which had high similarity to the nucleotide binding site (NBS) coding region of cereal cyst nematode resistance gene, Cre3， We designed three 3’ nested primers. Using single oligonucleotide nested PCR (SON-PCR) we successfully amplified one band, Rccn-L, of 1264bp from E-10 which is the wheat-Ae.variabilis translocation line containing the cereal cyst nematode resistance gene of Ae.variabilis. We found that this band of interesting is the 3’ flanking sequence of 1209bp in size of Rccn4. The coding region was 1026bp, which contained an incomplete open reading frame and a terminator codon, without initiation codon and intron, encoding a peptide of 342 amino acid residues, and shared 86﹪nucleotide sequence identity with Cre3. This peptide had a conserved LRR domain, containing the imperfect repeats,XXLXXLXXL, which contains 17﹪ leucine residues and shares, respectively, 89﹪ nucleotide sequence and 78﹪ amino acid sequence identity with the LRR sequence of Cre3 locus. This research firstly used SON-PCR in the research of plant genome successfully, which indicated that SON-PCR is another method of cloning plant gene. At the same time, According to the conversed motif of NBS and LRR region of cereal cyst nematode resistance gene Cre3 from wild wheat (Triticum tauschlii L.) and the known NBS-LRR group resistance genes, we designed two pairs of specific primers for NBS and LRR region respectively. One band of approximately 530bp was amplified using the specific primers for conversed NBS region and one band of approximately 1200bp was amplified with the specific primers for conversed LRR region. After sequencing, we found that these two sequences included 32bp common nucleotide sequence and have 1675 bp in total, which was registered as RCCN in the Genbank. RCCN contained a NBS-LRR domain and an incomplete open reading frame without initiation codon, terminator codon and inxon. Its exon encodes a peptide of 557 amino acid residues. The molecular weight of the protein from the amino acid was 63.537 KDa. The amino acid sequence of RCCN contained conserved motif: ILDD, ESKILVTTRSK, KGSPLAARTVGG, RRCFAYCS, EGF，LRR. RCCN shares 87.8﹪ nucleotide sequence and 77﹪ amino acid sequence identity with cereal cyst nematode gene Cre3. It might be a novel cereal cyst nematode resistance gene. These research results of cloning the resistance genes against cereal cyst nematode bring a great promise for transferring resistance genes into wheat cultivars and breeding new wheat varieties against cereal cyst nematode by gene engineering. And these results also lay the hard foundation for the expressing researches of these genes.

腹腔温热灌洗联合术后静脉微泵及置泵区域性动脉灌注化疗综合治疗胃肠道恶性肿瘤

目的:探讨胃肠道癌术后预防腹腔转移及肝转移的有效方法。方法:对157例胃肠道癌切除术后病人,随机分成术中腹腔温热灌洗及术后静脉微泵和置泵持续动脉灌注化疗组72例(简称治疗组),单纯静脉化疗组85例(简称对照组),并对其腹腔转移率、肝转移率及3年生存率进行对照研究。结果:治疗组腹腔转移率21.5%、肝脏转移率13.4%、3年生存率71.6%,对照组腹腔转移率41.2%、肝脏转移率22.6%、3年生存率47.5%。两组之间比较有显著性差异(P<0.05)。结论:术中腹腔温热灌洗及术后静脉微泵和置泵持续动脉灌注化疗,对胃肠道癌病人术后腹腔转移及肝转移有良好的防治作用。

Evolution of vegetation and climate since the last glacial maximum recorded at Dahu peat site；South China

IEECAS SKLLQG

黄土旱塬长期轮作施肥土壤剖面硝态氮的分布与积累

报道了黄土旱区连续 13年长期不同轮作施肥对土壤剖面硝态氮分布与积累的影响。化学氮肥的施用可有效地提高土壤硝态氮的含量 ,并造成硝态氮的淋溶 ,单施氮肥硝态氮淋溶深度达 150 cm;氮肥、磷肥和有机肥的配合施用不同程度的减小了硝态氮的淋溶 ,提高了氮肥的利用率 ;在贫氮地区土壤上 ,尤以氮肥、磷肥配施效果最佳。在相同的施肥种类和施肥量下 ,连续种植小麦、玉米、苜蓿 ,以玉米连作施肥土壤中硝态氮的累积量最小 ,而小麦连作施肥累积量最大 ;而且 ,无论施肥与否 ,均对深层土壤硝态氮分布造成不同程度的亏缺。在不同轮作系统中 ,粮草 3年轮作土壤剖面硝态氮的累积最小 ,其余轮作系统均造成土壤硝态氮不同程度的下淋积存

Nanoscale-enhanced Ru(bpy)(3)(2+) electrochemiluminescence labels and related aptamer-based biosensing system

A unique multilabeling at a single-site protocol of the Ru(bpy)(3)(2+) electrochemiluminescence (ECL) system is proposed. Nanoparticles (NPs) were used as assembly substrates to enrich ECL co-reactants of Ru(bpy)(3)(2+) to construct nanoscale-enhanced ECL labels. Two different kinds of NP substrates [including semiconductor NPs (CdTe) and noble metal NPs (gold)] capped with 2-(dimethylamino)ethanethiol (DMAET) [a tertiary amine derivative which is believed to be one of the most efficient of co-reactants of the Ru(bpy)(3)(2+) system] were synthesized through a simple one-pot synthesis method in aqueous media.

芸香苷铜(Ⅱ)络合物的电喷雾多级串联质谱分析

利用电喷雾离子阱质谱对黄酮苷类化合物———芸香苷与二价金属铜的络合物在软电离质谱条件下的络合行为进行了系统分析。所形成的 1∶1、2∶1、2∶2和 3∶2等不同化学计量比的芸香苷铜离子 (Ⅱ )络合产物 ,分别用 [R -H +Cu]+ ,[2R -H +Cu]+ ,[2R -3H +2Cu]+ 和 [3R -3H +2Cu]+ 表示。在多级串联质谱实验中 ,主要得到糖苷键断裂丢失寡糖链末端的鼠李糖基与苷元连接的葡萄糖基及整个寡糖链等中性碎片 ,同时也有寡糖链开环断裂和失水反应发生。

A new flow field and its two-dimension model for polymer electrolyte membrane fuel cells (PEMFCs)

A new flow field was designed to search flow fields fitting polymer electrolyte membrane fuel cells (PEMFCs) better due its extensible. There are many independent inlets and outlets in the new flow field. The new flow field we named NINO can extend to be more general when pressures at the inlet and outlet vary and some usual flow fields will be obtained. A new mathematical model whose view angle is obverse is used to describe the flow field.

Automorphism group algorithm in computer-aided structure elucidation

It is necessary to generate automorphism group of chemical graph in computer-aided structure eluciation. In this paper, an algorithm is developed by all-path topological symmetry algorithm to build automorphism group of chemical graph. A comparison of several topological symmetry algorithm reveals that all-path algorthm can yield correct of class of chemical graph. It lays a foundation for ESESOC system for computer-aided structure elucidation.

计算机辅助结构解析中自同构群的生成算法

在结构解析过程中，自同构群的生成是必须的。通过全通道拓朴等价性算法，得到了化合物的自同构群，并比较了几种拓扑等价性算法，结果发现，全通道算法能够正确划分化合物的结构图，从而为ＥＳＥＳＯＣ系统的立体异构体穷举生成奠定了基础。

末次冰期以来西赤道太平洋边缘海古气候变化研究

本文所研究的区域是西赤道太平洋的两个边缘海盆-苏禄海和南中国海。这两个边缘盆地具有明显不同的水文特征，南中国海底层水冷（<4 ℃）并且通气较好（溶解氧浓度>1.75ml/l），而苏禄海底层水温暖（>10 ℃）并且含氧量低（溶解氧浓度<1.25ml/l）。这些不同的水文特征导致了不同的沉积环境，同时两个海盆的沉积记录也包含了这些水文条件在不同地质时间上如何变化的历史。笔者分别对南中国海的GGC-6孔（12°09'1''E，118°03'9''，水深2775米）和苏禄海的GGC-29孔（8°17'7''E，118°49'9''N，水深1535米）的浮游和底栖有孔虫动物群进行了系统地分析鉴定和统计，并用转换函数对古温度进行了分析，力求通过有孔虫化石群的面貌和定量古生态分析来反映古气候的变化。通过对浮游有孔虫氧同位素环境效应的分析来探索本区的古气候变化。此外还对沉积物中的有机碳和碳酸钙的含量进行了测试和分析。通过上述工作我们得出以下几点认识：1、氧同位素和古水温记录可分末次冰期和全新世两期，由于岩心的长度所限地质记寻年代较短为24000年左右。2、浮游有孔虫G.Grassaformis的最后出现是苏禄海末次冰期极盛时（～18000B.P）重要的地质界线。3、发现在北欧地区短暂寒冷的仙女木期（Younger dryas）在本区也有发现。这在一定程度上说明了仙女木期存在的全球性。4、本区末次冰期与全新世的夏季表层古水温相差不到2 ℃与CLIMAP（1976）赤道太平洋末次冰期以来古水温度的模拟结果相附。冬季表层水温相差较大为3.5 ℃左右。5、瓷质壳底栖有孔虫的繁盛和翼足类的大量保存是苏禄海冰期时底层暖水的良好指示标志。6、本区有机碳的含量反映了底层水含氧浓度的变化。高有机碳代表低氧浓度，低有机碳代表高氧浓度。南中国海一直受到西北太平洋水体的影响，致使其底层水一直处于充氧状态，只是在冰期最盛时稍有降低。而苏禄海底层水氧浓度较低，尤其是末次冰期极盛时隔绝状态加强，大量的降水使表层海水淡化分层，致使底层水缺氧状态加剧。7、本区做为西赤道太平洋的边缘海，其CaCO_3旋回模式却与太平洋相反，而与大西洋一致，即冰期碳酸钙含量低，全新世冰后期碳酸钙含量高。冰期陆源物质供应的加强和全新世浮游有孔虫生产力的提高是控制本区碳酸钙旋回的主要因素。此外，冰期海水溶解作用的加强是南中国海冰期低CaCO_3含量的另一个重要原因，而苏禄海的碳酸钙沉积受海水溶解作用的影响却很小。8、浮游有孔虫N.dutertrei的大量增加和相对较轻的δ~(18)O值说明冰期最盛时海水表面的盐度较低。末次冰期时东南亚地区东北季风加强西南季风减弱，由于特殊的地理环境使本区在冰期时降水大量增加。

Emodin-induced apoptosis in human breast cancer BCap-37 cells through the mitochondrial signaling pathway

Emodin, a natural anthraquinone compound isolated from the rhizome of rhubarb, is reported to suppress the growth of tumor in many clinical situations. In this study, we focused on the effect of emodin in human breast cancer BCap-37 cells and further understand the underlying molecular mechanism in treating breast cancer. Using MTT assay and flow cytometry, we demonstrated the critical role of emodin in the suppression of the proliferation of BCap-37 cells based on a concentration- and time-dependent manner. The increase of apoptotic rate was also observed after incubation of BCap-37 cells on emodin at 20 mu M and 50 mu M for 48 h. The cells exhibited typical apoptotic features including cellular morphological change, chromatin condensation and membrane blebbing. The results of the study further showed that Bcl-2 level decreased, while Bax and cytosolic cytochrome c levels in sample cells increased after the emodin treatment by using Western blot. The decline in the Bcl-2/Bax ratio and the increase of cytosolic cytochrome c concentration were consistent with the increase of the apoptotic ratio. The results strongly suggest that the disruption of the mitochondrial signaling pathway was involved in emodin-induced apoptosis in BCap-37 cells.

基于模糊神经网络的多移动机器人自学习协调系统

研究多移动机器人的运动规划问题．针对机器人模型未知或不精确以及环境的动态变化，提出一种自学习模糊控制器（ＦＬＣ）来进行准确的速度跟踪．首先通过神经网络的学习训练构造ＦＬＣ，再由再励学习算法来在线调节ＦＬＣ的输出，以校正机器人运动状态，实现安全协调避撞

青岛近岸沉积物中持久性有机污染物多氯萘和多溴联苯醚

对同一样品经索氏抽提、多级复合硅胶层析柱、渗透凝胶层析柱、氧化铝层析柱分离, 利用低分辨率气相色谱-质谱方法对青岛近岸5个表层沉积物和1个贝类样品中的31种多氯萘(PCNs)和21种多溴联苯醚(PBDEs)异构体的含量分布特征、毒性当量浓度及来源进行了研究. 总PCNs和PBDEs最高含量出现在河口处. 总PCNs含量范围为212~1209 pg/g (dw), 以3氯代PCNs为主. 总PBDEs含量范围为117~5510 pg/g (dw). 主要以6-氯以下(含6-氯)为主. 局部来源的PCNs来自垃圾焚烧、燃煤等高温过程. PBDEs则主要来自分子扩散. 除了在河口处受到城市污水污泥的影响之外, 大气沉降是青岛近海PCNs与PBDEs的重要来源. 青岛近岸贻贝中PCNs和PBDEs都相对于沉积物有富集作用. PCNs的总毒性当量浓度均低于PCBs总毒性当量浓度.