花粉管通道遗传转化方法的新证据

将pBIN35S-mGFP4质粒转入受体菌DH5α中，扩繁后提取纯化质粒DNA，待棉花盛花期时，用花粉管通道转化方法将其注射到授粉24小时左右的子房中。在被检测的950个发育10～20天左右的幼胚中，发现七个幼胚在蓝光(460～490nm)激发下发出绿色荧光，而对照发出程度不同的红色荧光。将收获的“转化种子”浸泡萌发得到生长5～6天的黄化无菌苗，在200粒种子来源的无菌苗中，检出两棵转化植株。在紫外光照射灯下转化植株发出绿色荧光，其叶片和下胚轴横切面在蓝光激发下也与对照明显不同。PCR及Southern blotting结果均证实转化植株的真实性，从而为花粉管通道转化方法的可行性提供了直接可靠的细胞及分子生物学证据。 将GFPmut1质粒的gfp通过一端平接一端粘接后重组到pBI121的BamH1和Sal1限制性酶切位点从而代替GUS基因，然后将新的重组质粒用三亲交配法转入到LBA4404菌株中得到双元载体，用于棉花下胚轴切段的转化。结果表明，gfp象gus一样可作为报告基因用于农杆菌介导的棉花转化。在对筛选培养基上生长的“抗性愈伤”进一步进行报告基因检测时，只需手持紫外灯就可以检出GFP阳性愈伤，大大减少了工作量和试验费用。 另外，在进行农杆菌转化前的棉花卡那霉素敏感性实验中发现，卡那霉素对棉花下胚轴的愈伤组织形成和增殖均有明显的抑制作用，随其浓度的增加，愈伤组织形成的频率降低，增殖的倍数减小;当浓度增加到100mg/L时，愈伤组织严重褐化，其正常生长受到完全抑制;下胚轴形态学上端切段较下端更易受卡那霉素的影响。

玉米 rbcL 基因与兰藻 rbcS 基因定向融合及它们在宿主大肠杆菌中的表达

二磷酸核酮糖羧化酶/氧化酶(简称Rubisco, EC, 4.1.1.39)是绿色植物光合作用中参与固定CO2的关键酶。在高等植物，该酶是由8个分子量为55KD的大亚基(LSU)和8个分子量为14KD的小亚基(SSU)构成的16聚体。每个大亚基有四个活性中心，具有双向催化功能，其编码基因位于叶绿体基因组大单拷贝区;小亚基功能还不清楚，它由核基因组编码且有几个拷贝;未成熟小亚基N端有一段transit peptide，靠它的定向跨越叶绿体膜。迄今为止，取自几种植物材料的这二种亚基的氨基酸顺序和编码基因的核苷酸顺序分析业已完成。为该酶的遗传操作奠定了必要的基础。 由于Rubisco与人类利用太阳能和提高作物产量直接相关，所以成为通过生物技术进行改造的重大项目。 巢状假囊细菌(Anacyslis nidulans) R2是一种不含限制性内切酶的单细胞原核生物，能营光合作用，其Rubisco大亚基的氨基酸顺序与玉米的LSU同源性高达80%，但是第四个活性部位(Leu 456位)与玉米不同(Sys, 459位)，由此导致其对CO2的亲合力降低。另一方面，其rbcL与rbcS仅相隔93个bp，且同属一个操纵子。这意味着有可能用同源DNA片段等位交换的办法来改造其rbcL基因。 根据现有的资料，设计出玉米rbcL与兰藻rbcS定向重组于pUC119的兰图：先从pANP1155中切出0.7kb含蓝藻rbcS的PstI-HindIII片段，克隆进pUC119的lacZ启动子下游得pTAS28，采用Reverse primer作引物进行核苷酸顺序分析，确认蓝藻rbcS基因座落在pTAS28正链上。随后从pZmc460中切出包含玉米rbcL基因1.7kb的BglII-HincII片段，将它插入pTAS28的HincⅡ-BamHⅠ双酶切位点，得到pTMN3;为了比较，在另一个质粒pTMN7于1.7kb片段之前加进0.1kb的PstI-HaeIII蓝藻DNA。根据玉米rbcL基因核苷酸顺序（1218-1251）合成一个Oligonucleotide probe，对这三个质粒的总RNA抽提物进行Northern Blot，得到明显的杂交斑点;接着用菌体总蛋白冻干品进行了Western分析，并以新鲜的玉米和烟草叶片为对照，得到阳性结果。显然这二种基因重组之后仍能在宿主E. coli中正常表达。 真正的挑战应是下一步用上述二种质粒转化兰藻A. nidulans R2，考查其能否整合进基因组并表现出较低的氧化酶活性。

大蹼铃蟾抗菌肽- 2 与IFN2α22b 和TNF2α对膀胱癌细胞株抑制活性的实验研究

目的:比较大蹼铃蟾抗菌肽- 2 (Maximin 2) 与肿瘤坏死因子2α(TNF2α) 、干扰素2α22b ( IFN2α22b) 对不同膀胱癌细胞株的抑制杀伤作用。方法:采用细胞培养及MTT 比色法,检测Maximin 2 、TNF2α及 IFN2α22b 对3 个不同膀胱癌细胞株的抑制作用及作用特点,计算量效回归曲线及50 %抑制率。结果: Maximin 2 对不同的膀胱癌细胞株在100μgPmL 左右(48 h) 即达到50 %抑制率浓度( IC50) ,并呈剂量相 关性,与TNF2α、IFN2α22b 抑制作用的表现形式不同。Maximin 2 抑制率最高表现在第48 h ,以后逐渐减 弱, TNF2α、IFN2α22b 则在前24 h 作用较弱,以后逐渐增强。结论:Maximin 2 对不同的膀胱癌细胞株均 有较强的抑制作用,同TNF2α、IFN2α22b 比较其作用机制有所不同,Maximin 2 对膀胱癌细胞株的杀伤作 用可能主要是通过对肿瘤细胞的直接抑制,并且相对容易失活。

Two antimicrobial peptides from skin secretions of Rana grahami

Two antimicrobial peptides manifested a broad spectrum of anti microbial activity against various microorganisms have been isolated from skin secretions of Rana grahami. These antimicrobial peptides were named grahamin 1 and grahamin 2. Their primary Stru

马来熊的 DNA 序列分析与遗传多样性研究

马来熊在 IUCN 红皮书中被列为受胁动物, 其保护受到广泛的关注. 该文研究４只马来熊的线粒体 DNA 序列, 其中１只来自云南, 其余３只产地不详, 但来自不同的搜集渠道. 对于每个个体, 作者测定了 397bp 的细胞色素ｂ基因、346bp 的12S rRNA基因、98bp 的 tRNA 基因和333bp 的Ｄ环区序列, 共计1174bp. 经与黑犀牛序列比较, 发现 RNA 基因的空间结构对基因的进化有显著影响, 环区的进化明显快于茎区的. 对于细胞色素ｂ基因、12S rRNA基因和 tRNA 基因, 在马来熊个体间未发现序列变异. 这一结果提示, 马来熊群体的遗传变异程度低. 在D环区, 有13和1个位点分别出现转换和颠换. 根据D环区序列, 作者采用简约法确定了马来熊群体间的进化关系. 作者的结果表明, 线粒体 DNA 的D环区是研究马来熊群体遗传结构十分有效的遗传结构十分有效的遗传标记。

Analysis of 18S rRNA gene of Octostigma sinensis (Projapygoidea : Octostigmatidae) supports the monophyly of Diplura

The phylogenetic position of Diplura within Hexapoda has been controversial. There are three major lineages in Diplura: Campodeoidea, Projapygoidea, and Japygoidea. However, most of the previous studies were restricted to Campodeoidea and Japygoidea. Unti

代谢途径的进化

生物的新陈代谢是通过一系列的代谢途径来实现的.这些调节精妙、相瓦协作的代谢途径是如何进化形成的一直是一个引人入胜的重要问题.自1945年有关该问题的第一个假说--"逆向进化假说"提出以来,迄今已发展出七种假说,包括"逆向进化假说"、"半酶理论"、"向前发展模型"、"酶的招募假说"、"多功能酶特化假说"、"整个代谢途径的复制假说"和"从头创造假说".其中最受关注的是"逆向进化假说"和"酶的招募假说",而最近提出的"多功能酶特化假说"由于有较好的理论基础和实验证据支持,也逐渐引起人们的关注.本文对这些假说逐一作了概述,并结合作者的相关研究工作,对该领域的研究现状和发展趋势进行了分析讨论和展望.

柑橘凤蝶生殖系统解剖学研究

通过解剖柑橘凤蝶成虫的生殖系统，研究了柑橘凤蝶生殖器官的形态及结构特点。

Intergroup interactions in Tibetan macaques at Mt. Emei, China

Data on intergroup-interactions (I-I) were collected in 5 seasonally provisioned groups (A, B, D, D-1, and E) of Tibetan macaques (Macaca Thibetana) at Mt. Emei in three 70-day periods between 1991 April-June (P1), September-November (P2), December-1992 February (P3). The I-I were categorized as forewarning made by high-ranking males (including Branch Shaking and/or Loud Calls), long-distance interactions in space (specified by changes in their foraging movements), and close encounters (with Affinitive Behavior, Male's Herding Female, Sexual Interaction, Severe Conflict, Adult Male-male Conflict, Opportunistic Advance and Retreat, etc. performed by different age-sex classes). From periods Fl to P3, the I-I rate decreased with reduction in population density as a positive correlate of food clumpedness or the number of potential feeders along a pedestrian trail. On the other hand, from the birth season (BS, represented by P1 and P3) to the mating season (MS, represented by P2) the dominance relation between groups, which produced a winner and a loser in the encounters, became obscure; the proportion of close encounters in the I-I increased; the asymmetry (local groups over intruders) of forewarning signals disappeared; the rate of branch shaking decreased; and sometimes intergroup cohesion appeared. Considering that sexual interactions also occurred between the encountering groups, above changes in intergroup behaviors may be explained with a model of the way in which the competition for food (exclusion) and the sexual attractiveness between opposite sexes were in a dynamic equilibrium among the groups, with the former outweighing the latter in the BS, and conversely in the MS. Females made 93% of severe conflicts, which occurred in 18% of close encounters. Groups fissioned in the recent past shared the same home range, and showed the highest hostility to each other by females. In conspicuous contrast with females' great interest in intergroup food/range competition, adult male-male conflicts that were normally without body contact occurred in 66% bf close encounters; high-ranking male herding of females, which is typical in baboons, appeared in 83% of close encounters, and showed no changes with season and sexual weight-dimorphism; peripheral juvenile and subadult males were the main performers of the affinitive behaviors, opportunistic advance and retreat, and guarding at the border. In brief, all males appeared to "sit on the fence" at the border, likely holding out hope of gaining the favor of females both within and outside the group. Thus, females and males attempted to maximize reproductive values in different ways, just as expected by Darwin-Trivers' theory of sexual selection. In addition, group fission was observed in the largest and highest-ranking group for two times (both in the MS) when its size increased to a certain level, and the mother group kept their dominant position in size and rank among the groups that might encounter, suggesting that fission takes a way of discarding the "superfluous part" in order to balance the cost of competition for food and mates within a group, and the benefit of cooperation to access the resources for animals in the mother group. (C) 1997 Wiley-Liss, Inc.

许氏平鲉幼鱼的赖氨酸需求量

通过8周的生长实验确定了许氏平(Sebastes schlegeli)幼鱼的赖氨酸需求量。饲料必需氨基酸组成(除赖氨酸外)参照鱼体肌肉蛋白的氨基酸模式,配制赖氨酸含量为1.542、.04、2.54、3.04、3.54和4.04%的等氮等能的六种半精制饲料。研究结果表明,随着饲料中赖氨酸含量的增加,增重率逐渐提高,当饲料中赖氨酸含量为3.54%时,达到最大值(P<0.05)。基础饲料组的蛋白质贮积率最低,且显著低于3.54%组(P<0.05),和其他组之间差异不显著(P>0.05)。随着饲料中赖氨酸含量的增