激素联合丙种球蛋白治疗小儿重症肌无力的疗效及对患儿免疫球蛋白和补体的影响

目的：探讨激素联合丙种球蛋白治疗小儿重症肌无力的临床疗效及对患儿免疫球蛋白和补体的影响。方法：回顾性分析在我院治疗的70例重症肌无力患儿的临床资料,采用随机序号的方式将其分为观察组和对照组各35例,观察组给予甲泼尼龙联合丙种球蛋白,对照组仅给予甲泼尼龙,观察两组的临床疗效及免疫球蛋白和补体变化情况。结果：观察组总有效率为94．3％明显优于对照组74．3％,两组比较有统计学意义（P〈0．05）;观察组症状明显缓解时间（6．55±1．35）d以及总住院天数（17．15±3．65）d较对照组明显缩短,两组比较差异均有统计学意义（P〈0．05）。结论：采用激素联合丙种球蛋白治疗小儿重症肌无力,可以明显改善患者肌无力症状,获得较为满意的临床疗效,值得进一步推广使用。     

低温胁迫对灰飞虱种群生长发育与繁殖的影响

为了探讨低温在灰飞虱Laodelphax striatellus(Fallén)种群发生发展中的作用,本文以南京冬季的旬平均最低温为依据,设定出-6～2℃的低温,研究了用该低温处理3～4龄若虫不同时间后,灰飞虱的存活、生长与繁殖特性。结果表明,灰飞虱3～4龄若虫有较强的耐低温能力,-6～2℃的温度下,其半致迷和半致死时间分别为1.22和20.49d。低温抑制了若虫的生长发育,发育历期明显延长。低温处理后成虫的性比有所提高,并且短翅率显著增加。3～4龄若虫低温处理1～7d后羽化的成虫有产卵前期缩短、寿命延长的趋势,但对繁殖力没有显著影响,并且低温处理2d后的产卵量要稍高于没有处理的对照。由此可见,-6～2℃低温的短期胁迫对灰飞虱种群的发生发展不会产生抑制作用。     

用DREAM技术纠正人工合成基因中的突变

目的：介绍一种简便、有效的定点突变技术。方法：根据突变位点附近的DNA序列推导出氨基酸序列,再以此氨基酸序列进行逆翻译,这样在不改变氨基酸序列的前提下可以得到数目巨大的隐性突变体（silent mutants）,这些突变体中包含大量的限制性内切酶位点,选择合适的酶切位点设计引物用PCR技术扩增两侧DNA片段,然后以相应酶切融合这两个片段即可完成定点突变。结果：用该方法成功地在人工合成的含有缺失的可溶性组织因子基因的472位插入C,T两个碱基,校正了阅读框架,获得了预期的目的基因。结论：该方法简便、有效, 避免了多轮PCR和合成长引物导致突变的可能性,这种改进的PCR 定点诱变技术我们称之为“设计限制酶辅助突变”（Designed Restriction Enzyme Assisted Mutagenesis, DREAM）。此技术简单方便, 诱变的成功率高, 适于实验室常规应用。     

竹红菌甲素在脂质体中的光谱性质和结合能力研究

竹红菌甲素在脂质体中的光谱性质和结合能力研究邹伟,安静仪,蒋丽金（中国科学院北京感光化学研究所,１００１０１）关键词竹红菌甲素;光谱特性;结合;脂质体竹红菌甲素（Ｒ人）是一种新型并配类光疗药物,临床上治疗一些皮肤病效果显著”’,研究表明ＨＡ对癌细胞有...     

猪繁殖与呼吸综合征病毒ORF2～4特异siRNA的筛选及其抑制病毒复制效果的研究

RNA干扰(RNAi)技术是基因功能研究的有效工具,为了了解猪繁殖与呼吸综合征病毒(PRRSV)次要结构蛋白GP2、GP3、GP4在病毒复制中的作用,针对各自的编码基因ORF2、ORF3、ORF4分别选取4个小干扰RNA(siRNA)位点(共12个),构建相应的短发夹RNA(shRNA)表达载体,转染MARC-145细胞后,通过荧光定量PCR和病毒滴度检测干扰效果。筛选了可以减少GP2、GP3、GP4相应基因mRNA含量的ORF2、ORF3、ORF4特异shRNA表达载体,病毒效价滴定表明shRNA表达载体处理细胞可以减少GP2、GP3、GP4相应基因mRNA含量,细胞培养上清中的病毒滴度比对照低184~4.65倍。     

The auxin response factor,OsARF19, controls rice leaf angles through positively regulating OsGH3‐5 and OsBRI1

Auxin and brassinosteroid (BR) are important phytohormones for controlling lamina inclination implicated in plant architecture and grain yield. But the molecular mechanism of auxin and BR crosstalk for regulating lamina inclination remains unknown. Auxin response factors (ARFs) control various aspects of plant growth and development. We here report that OsARF19‐overexpression rice lines show an enlarged lamina inclination due to increase of its adaxial cell division. OsARF19 is expressed in various organs including lamina joint and strongly induced by auxin and BR. Chromatin immunoprecipitation (ChIP) and yeast one‐hybrid assays demonstrate that OsARF19 binds to the promoter of OsGH3‐5 and brassinosteroid insensitive 1 (OsBRI1) directing their expression. OsGH3‐5‐overexpression lines show a similar phenotype as OsARF19‐O1. Free auxin contents in the lamina joint of OsGH3‐5‐O1 or OsARF19‐O1 are reduced. OsGH3‐5 is localized at the endoplasmic retieulum (ER) matching reduction of the free auxin contents in OsGH3‐5‐O1. osarf19‐TDNA and osgh3‐5‐Tos17 mutants without erected leaves show a function redundancy with other members of their gene family. OsARF19‐overexpression lines are sensitive to exogenous BR treatment and alter the expressions of genes related to BR signalling. These findings provide novel insights into auxin and BR signalling, and might have significant implications for improving plant architecture of monocot crops.     

基于表面等离子体共振和电化学联用的DNA传感器研究进展

表面等离子共振(surface plasmon resonance,SPR)技术旨在检测物体表面附近折射率的变化,其特点是无标记、实时、灵敏和快速,该技术多用于研究分子的相互作用,包括动力学、效率常数和大分子构象变化等。电化学(electrochemical,EC)技术是一项用于定性定量研究电子转移、物质氧化还原、界面吸附等过程的成熟技术,具有简单、低成本和设备小型化的优点。现有的DNA杂交技术,例如光学、电化学或压电转导技术,主要关注于提高DNA杂交检测系统的选择性和灵敏度。传统的SPR在DNA分析方面,由于无法测量折射率的极小变化而在超灵敏检测中的应用受到限制。因此,随着纳米材料的研发和联用技术的飞速发展,SPR与EC联用的生物传感器研究越来越成为人们关注的热点。近年来,关于SPR和EC联用在DNA检测方面的综述鲜有报道。对SPR和EC检测DNA的技术原理、联用方法、应用进展等方面作出了简要的介绍,以期为表面等离子共振和电化学联用的DNA传感器相关研究提供参考。     

Uncovering species boundaries in the Neotropical ant complex Ectatomma ruidum (Ectatomminae) under the presence of nuclear mitochondrial paralogues

Nuclear mitochondrial (mt) paralogues (numts) are non‐functional fragments of mtDNA integrated into the nuclear genome that can overestimate the number of species in analyses based on mtDNA sequences. As numts have relatively slow mutation rates, they can pass undetected by conventional procedures such as inspecting for internal stop codons, indels or apparent polymorphism in chromatograms. Species boundaries based on mtDNA markers therefore require a thorough assessment of numts, especially in insects, where this phenomenon appears to be relatively frequent. Ectatomma ruidum is a widely distributed Neotropical ant species that is distributed from northern Mexico to northern Brazil. Previous behavioural and molecular evidence suggests that this species actually represents a composite taxon. Here we assessed the species boundaries in E. ruidum based on two mt (COI, cyt b) and one nuclear (H3) marker, as well as on external morphology. Ancient and recent mt paralogues were detected in several specimens, although pre‐PCR dilution of DNA template helped to recover most of the mt orthologues. Based on the congruence found between our species delineation obtained from the mt genealogies and the discriminated morphospecies, we propose that E. ruidum is actually composed of at least three species. Two of these species have a wide geographical distribution in the Neotropics, whereas the remaining one was restricted to localities situated near the Pacific coast in south‐east Mexico. We also found extensive intra‐ and interspecific variation in the barcoding locus. Moreover, the nuclear evidence suggests the existence of hybrids between two of these species in Oaxaca, south‐east Mexico. This study agrees with previous studies of other closely related animal taxa, which have revealed a complex evolutionary history and overlooked species diversity in the latter region.     

双光子及共聚焦激光扫描显微术研究天花粉蛋白对人绒癌细胞的作用机制

天花粉蛋白(trichosanthin, TCS)是从中草药栝楼根中提取的一种核糖体失活蛋白,具有抗肿瘤和抗HIV功能.应用双光子及共聚焦激光扫描显微术结合特异性荧光探针Hoechst 33342、2′,7′-二氯荧光黄双乙酸酯 (DCFH-DA)、Indo-1和Fluo 3-AM,首次同时观察了TCS诱导人绒癌细胞（JAR细胞）凋亡过程中活性氧自由基（ROS）和细胞内钙离子浓度（［Ca²⁺］_i）的变化,实验结果表明TCS引起的［Ca²⁺］_i升高和ROS形成参与了TCS诱导的JAR细胞凋亡,并且ROS形成和［Ca²⁺］_i升高有关.共聚焦激光扫描显微术的研究结果表明,［Ca²⁺］_i升高不是导致ROS形成的主要原因,TCS诱导产生的ROS可能是通过TCS与JAR细胞膜表面受体作用介导的.     

一种利用Cre/loxP系统进行标记基因删除与靶基因置换的细胞模型研究

Cre/lox系统可以介导DNA的定点插入和定点删除,可利用其实现转基因动物中"友好位点"的重复利用及标记基因的有效删除.为直观地评估该系统介导的以上两种重组反应的效果,通过标记基因并利用大鼠乳腺癌细胞系SHZ-88进行了模型研究.首先构建了两个载体:a.整合载体pTE-lox2272-DsRed-loxP-GFP-loxP,含有红色荧光标记基因DsRed和绿色荧光标记基因GFP;b.置换载体pT-lox2272-neo-loxP,含有筛选标记基因neo,用以置换DsRed基因.然后,用整合载体转染SHZ-88细胞,并随机挑取了3个同时表达DsRed和GFP的稳定整合细胞克隆.随后用置换载体和Cre表达载体PBS185对以上每个克隆分别进行了3次共转染,通过G418筛选并扩增培养后,总共获得1 070个克隆.通过分析标记基因DsRed和GFP在这些克隆中的表达情况:Cre介导的删除效率为91.1%,定点置换效率为29.3%.最后对部分克隆进行了PCR和DNA印迹鉴定,分子鉴定结果与发光的表型状况一致.这一方法为Cre/lox系统在转基因家畜生产中的进一步应用提供了实验依据.