油菜profilin基因的克隆和表达分析

profilin是高等植物中的一种与肌动蛋白结合的蛋白,采用RT-PCR技术克隆了油菜（Brassica napus L.cv.canadian tween)花粉中的一个369bp的cDNA片段,序列分析结果表明,该cDNA与已报道的其他植物的profilin基因具有较高核酸序列同源性,与玉米（Zea maysL.)基因同源性为82%,拟南芥（Arabidopsis)基因同源性为85%,水稻（Oryza sativa L.)基因同源性为81%,烟草（Nicotiana tabacum L.)基因同源性为82%,结论5′RACE和3′RACE技术,获得了全长cDNA,其为672bp。该cDNA包含一个开放密码框。5′未翻译区和一个带有Poly(A)的3′区域。Northern杂交结果显示它主要在花粉和花药中表达。     

棉花PTS2受体基因(GhPex7)的克隆及表达分析

利用cDNA—AFLP差异片段F010,通过RACE延伸、EST、检索等方法获得了一个棉花过氧化物酶体定位信号2受体蛋白基因(peroxisomal targetingsignal type 2 receptor,GhPex7p)的编码序列。该cDNA包含一个954bp的开放阅读框,编码317个氨基酸,推测其等电点为5．603。同源性分析表明：推测GhPex7与拟南芥、酵母、果蝇、小鼠和人的Pex7p基因存在序列相似性,其中与拟南芥的同源性最高,为83％,并具有3段WD-40蛋白家族的保守域,与拟南芥AtPex7的编码蛋白同类。Southern杂交结果表明该基因在陆地棉基因组中存在两个拷贝。Northern blotting和RT-PCR分析表明该基因在棉花根、茎、叶、花、胚珠和纤维中均表达,但茎、叶组织中的表达水平明显高于胚珠和纤维。     

棕色棉F3'H-羟化酶基因的克隆与生物信息学分析

根据葡萄的类黄酮3′-羟化酶(F3'H)基因全长cDNA序列Blast所得棉花的EST序列设计引物,以开花后16 d(DPA16)的新彩棉5号(xC-5)纤维为材料,利用RACE和RT-PCR技术分离得到了2个类黄酮3′-羟化酶基因cDNA序列,此2个序列编码区完全相同,仅在3'UTR区存在片段长短的差异,推测可能是基因转录后加工方式不同所造成.克隆所获得的棉花F3'H基因编码区全长1 533 bp,编码510个氨基酸,氨基酸序列分析预测表明,该基因所编码蛋白含有一个跨膜结构域,是一种分泌蛋白,定位于内质网上,并含有一段与细胞色素P450功能区相匹配的保守功能域;序列比对结果表明,棉花F3'H基因与其他多个物种的F3'H基因在氨基酸序列上有较高的同源性;聚类分析结果表明,棉花F3'H蛋白与双子叶植物大豆的F3'H亲缘关系较为接近,而与单子叶植物高梁等作物则较远.     

鸡二价金属转运蛋白1 cDNA的克隆与序列分析

鸡二价金属转运蛋白1（divalent metal transporter 1, DMT1）在动物胃肠道锰吸收过程中起重要作用.根据哺乳动物Dmt1同源蛋白氨基酸序列的保守性设计引物,应用3′RACE(rapid amplification of cDNA ends)技术,扩增并克隆获得鸡小肠Dmt1 cDNA 3′端1 289bp和1 092bp的2种片段,发现其3′端翻译区和非翻译区存在差异. 根据鸡Dmt1 cDNA 3′端片段的测序结果设计引物,扩增获得1个与3′端片段部分重叠的鸡Dmt1 cDNA 5′端907 bp片段,并对其进行了克隆测序. 根据鸡小肠Dmt1 3′RACE片段和5′RACE片段序列信息进行拼接,从而获得鸡小肠Dmt1 cDNA全序列信息.结果表明,鸡小肠Dmt1 cDNA有2种形式,1种全长为1 972个核苷酸,其中5′非翻译区为104个核苷酸,编码区1 695个核苷酸,3′非翻译区为173个核苷酸,编码1个含564个氨基酸残基的蛋白质;另1种形式为1 775个核苷酸,其中5′非翻译区为104个核苷酸,编码区1 593个核苷酸,3′非翻译区为78个核苷酸,编码1个含530个氨基酸残基的蛋白质.据鸡Dmt1 cDNA推测出的2种形式蛋白质的氨基酸序列与人、大鼠和小鼠的Dmt1蛋白具有高度同源性,它们的同源性分别为82%、82%、80%,和 84%、84%、83%. 对推测氨基酸序列进行疏水性和跨膜区分析表明,Dmt1蛋白为1种跨膜整合蛋白,具有膜转运蛋白糖基化位点和底物结合位点的保守序列.     

一种苦荞主要过敏原基因cDNA的克隆及序列分析

为了获得苦荞中主要过敏原的cDNA和由此推导的蛋白质序列 ,分析其结构特点 ,以苦荞幼根根尖为材料 ,提取总RNA并反转录mRNA为cDNA第一链 .通过RT PCR、3′RACE、基因克隆及序列测定 ,获得一种苦荞主要过敏蛋白基因的cDNA片段 (GenBank登录号为AY0 4 4918) .该cDNA片段由 76 8bp组成 ,包括 3′端非编码区 180个bp ,开放阅读框 5 88bp .可编码一个由 195个氨基酸残基组成的功能蛋白及一个终止密码 .苦荞主要过敏原基因与甜荞 2 2kD过敏蛋白、豆球类蛋白的核苷酸序列分别有 95 %和 93%的同源性 .其推导的氨基酸序列与甜荞球蛋白、刀豆蛋白、甜橙柠檬素分别有 93%、83%和 5 7%的同源性 .该过敏蛋白 183～ 188位氨基酸残基KEEEKE在多数不同过敏原中均存在 ,推测可能为其中的抗原决定簇序列     

彩叶草中一个富亮氨酸重复相关蛋白基因SsLRP的克隆和分析

根据彩叶草叶片小型EST库中一条具有1个富亮氨酸重复（leucine-rich repeat,LRR）结构域的EST序列,采用RACE与文库结合的方法,克隆了1个具有5个LRR结构域的全长cDNA,SsLRP（LRR-Related Protein）（GenBank登录号FJ787729）。SsLRP cDNA全长1024bp,包含一个657bp的ORF框,编码218个氨基酸。其5’-UTR区含有2个终止子TAG,3’-UTR区具有推测的加尾信号AATAAA。SsLRP蛋白N端具有的信号肽和保守的亮氨酸拉链结构域,具有5个保守的LRR结构域,多个磷酸化位点和N-糖基化位点。多序列比对和系统进化分析表明,SsLRP与番茄SlLRP同源性最高。二级结构和三级结构预测表明,SsLRP的功能可能与保守的LRR结构域密切相关,推测该基因可能参与蛋白间的相互作用与信号识别。RT-PCR分析表明,SsLRP与番茄SlLRP具有相似的表达模式,在正常植株的根、茎、叶和花中都有表达,在受菌核病感染植株的茎和叶中表达上调。     

天麻中一种抗真菌蛋白基因的克隆

依据天麻(GastrodiaelataBl.f.flavidaS.Chow)抗真菌蛋白GAFP_1的N端部分氨基酸序列设计简并引物,通过RACE(快速分离cDNA末端)的方法扩增得到GAFP_1全长cDNA。该cDNA包含一个编码171个氨基酸的ORF,推导的多肽序列与测得的蛋白质部分序列相同;在5′端有一个长为55bp的5′非编码区;终止密码子下游有一个141bp长的3′非编码区,其中含有两个加poly(A)信号及长度为26个腺苷酸的poly(A)。经检索发现该推导蛋白序列与火烧兰(Epipactishelloborine)和二叶兰(Listeraovata)的甘露糖结合蛋白以及雪花莲(Galanthusnivalis)中的甘露糖结合凝集素具有很高同源性。     

小麦抗黄矮病基因相关cDNA片段的克隆

以从小麦品种间杂交组合川35050/山农483 F7株系中分离出的感、抗小麦黄矮病的2个近等基因系为材料,利用DDRT-PCR技术和抗病基因保守结构域序列设计的7条简并或特异引物进行差异显示,PCR分析获得4条差异序列,并分别命名为:Rbdv1~Rbdv4(GenBank注册号:EU267934~EU267937).以Rbdv1为靶序列,利用RACE对其3′末端进行PCR扩增,得到长778 bp、末端有poly(A)尾巴的序列.用DNAMAN软件将3′RACE得到的序列与靶序列拼接得到1 196 bp长的片段,命名为A1(GenBank注册号:EU267938).BlastN分析表明,A1与拟南芥、水稻中CDC48蛋白的同源性分别为81%和90%,其氨基酸序列中包含1个P-Loop,具有R基因的特征结构域,推测该片段很可能与抗小麦黄矮病基因相关.     

一种新的甘露糖结合凝集素--朱顶兰凝集素基因的克隆及序列分析

运用同源克隆的方法设计简并引物,通过3′和5′RACE技术,从石蒜科植物朱顶兰(Amaryllis vittata Ait)总RNA中克隆了编码此凝集素(AVA)的全长cDNA序列.该基因全长686 bp,起始密码子位于第41～43 bp,终止密码子位于515～517 bp处,开放阅读框长474 bp,编码158个氨基酸,包含信号肽序列、成熟蛋白序列和C-末端剪切序列的前体蛋白.成熟蛋白由109个氨基酸残基组成,分子量为11.9kD.成熟蛋白在氨基酸水平上与雪花莲凝集素、水仙凝集素、石蒜凝集素和君子兰凝集素分别有73.4%、85.3%、80.7%和83.5%的同源性;朱顶兰凝集素的分子模式显示其与雪花莲凝集素有极其相似的三维结构;在Blocks数据库中检索AVA蛋白氨基酸序列的结构域,发现有3个凝集素功能结构域,并具有3个典型的甘露糖专一结合位点盒(QDNY).     

鲢鱼解偶联蛋白2全长cDNA序列的克隆及其组织表达

从淡水食毒藻鱼类鲢鱼(Hypophthalmichthysmolitrix)肝脏,通过简并引物克隆解偶联蛋白2(un-couplingprotein2,UCP2)cDNA核心序列,应用5′RACE和3′RACE技术分别扩增该序列的5′末端和3′末端序列,最后通过序列拼接获得鲢鱼肝脏UCP2cDNA全序列。序列分析结果表明,鲢鱼肝脏UCP2cDNA全长1452bp,其中5′-UTR长337bp,3′-UTR长182bp,编码区933bp,编码310个氨基酸,推测的氨基酸序列包含线粒体内膜载体蛋白3个特征结构及解偶联蛋白(UCPs)的特征序列。对鲢鱼不同组织UCP2的表达调控研究发现,鲢鱼组织UCP2基因在肠道、肝脏、肌肉、脂肪组织均大量表达,而在脑组织表达量较低,这与鲢鱼体内微囊藻毒素在这几个组织的分布完全一致,表明UCP2的功能可能与抑制微囊藻毒素引发过量活性氧(ROS)生成有关。     

Physicochemical and biological properties of 6(1),6(3),6(5)-tri-O-alpha-maltosyl-cyclomaltoheptaose (6(1),6(3),6(5)-tri-O-alpha-maltosyl-beta-cyclodextrin)

A unique multibranched cyclomaltooligosaccharide (cyclodextrin, CD) of 6(1),6(3),6(5)-tri-O-alpha-maltosyl-cyclomaltoheptaose [6(1),6(3),6(5)-tri-O-alpha-maltosyl-beta-cyclodextrin, (G(2))(3)-betaCD] was prepared. The physicochemical and biological properties of (G(2))(3)-betaCD were determined together with those of monobranched CDs (6-O-alpha-D-glucopyranosyl-alpha-cyclodextrin (G(1)-alphaCD), 6-O-alpha-D-glucopyranosyl-beta-cyclodextrin (G(1)-betaCD), and 6-O-alpha-maltosyl-beta-cyclodextrin (G(2)-betaCD)). NMR spectra of (G(2))(3)-betaCD were measured using various 2D NMR techniques. The solubility of (G(2))(3)-betaCD in water and MeOH-water solutions was extremely high in comparison with nonbranched betaCD and was about the same as that of the other monobranched betaCDs. The formation of an inclusion complex of (G(2))(3)-betaCD with stereoisomers (estradiol, retinoic acid, quinine, citral, and glycyrrhetinic acid) depends on the cis-trans isomers of guest compounds. The cis isomers of estradiol, retinoic acid, and glycyrrhetinic acid were included more than their trans isomers, while the trans isomers of citral and quinine fit more tightly than their cis isomers. (G(2))(3)-betaCD was the most effective host compound in the cis-trans resolution of glycyrrhetinic acid. Among the branched betaCDs, (G(2))(3)-betaCD exhibited the weakest hemolytic activity in human erythrocytes and showed negligible cytotoxicity in Caco-2 cells up to 200 microM. These results indicate unique characteristics of (G(2))(3)-betaCD in some biological responses of cultured cells.     

Spectral and thermodynamic properties of Ag(I), Au(III), Cd(II), Co(II), Fe(III), Hg(II), Mn(II), Ni(II), Pb(II), U(IV), and Zn(II) binding by methanobactin from Methylosinus trichosporium OB3b

Methanobactin (mb) is a novel chromopeptide that appears to function as the extracellular component of a copper acquisition system in methanotrophic bacteria. To examine this potential physiological role, and to distinguish it from iron binding siderophores, the spectral (UV–visible absorption, circular dichroism, fluorescence, and X-ray photoelectron) and thermodynamic properties of metal binding by mb were examined. In the absence of Cu(II) or Cu(I), mb will bind Ag(I), Au(III), Co(II), Cd(II), Fe(III), Hg(II), Mn(II), Ni(II), Pb(II), U(VI), or Zn(II), but not Ba(II), Ca(II), La(II), Mg(II), and Sr(II). The results suggest metals such as Ag(I), Au(III), Hg(II), Pb(II) and possibly U(VI) are bound by a mechanism similar to Cu, whereas the coordination of Co(II), Cd(II), Fe(III), Mn(II), Ni(II) and Zn(II) by mb differs from Cu(II). Consistent with its role as a copper-binding compound or chalkophore, the binding constants of all the metals examined were less than those observed with Cu(II) and copper displaced other metals except Ag(I) and Au(III) bound to mb. However, the binding of different metals by mb suggests that methanotrophic activity also may play a role in either the solubilization or immobilization of many metals in situ.     

Circular dichroism spectra of DNA quadruplexes [d(G(5)T(5))](4) as formed with G(4) and T(4) tetrads and [d(G(5)T(5)). d(A(5)C(5))]2 as formed with Watson-Crick-like (G-C)(2) and (T-A)(2) tetrads

We utilize electrophoresis and find that a thermally treated equimolar mixture of the oligonucleotide d(G(5)T(5)) and its complementary oligonucleotide d(A(5)C(5)) exhibits either two bands or a single band in one lane, depending on the conditions of the incubation solutions. The thermally treated d(G(5)T(5)) solution loaded in a different lane exhibits a single band of the parallel quadruplex [d(G(5)T(5))](4), which is composed of homocyclic hydrogen-bonded G(4) and T(4) tetrads previously proposed. For the thermally treated equimolar mixture of d(G(5)T(5)) and d(A(5)C(5)), the fast band is assigned to a Watson-Crick d(G(5)T(5)). d(A(5)C(5)) duplex, so that the slow band with the same low mobility as that of [d(G(5)T(5))](4) may be assigned to either [d(G(5)T(5))](4) itself or a [d(G(5)T(5)). d(A(5)C(5))](2) quadruplex. If the latter compound is true, this may be the antiparallel quadruplex composed of the heterocyclic hydrogen-bonded G-C-G-C and T-A-T-A tetrads proposed previously. After removing these three bands for the duplex and two kinds of hypothetical quadruplexes, we electrophoretically elute the corresponding compounds in the same electrophoresis buffer using an electroeluter. The eluted compounds are ascertained to be stable by electrophoresis. The circular dichroism (CD) and UV absorption spectra measured for the three isolated compounds are found to be clearly different. For the electrophoretic elution of the hypothetical [d(G(5)T(5))](4) quadruplex, the result of the molecularity of n = 4 obtained from the CD melting curve analysis provides further support for the formation of the parallel [d(G(5)T(5))](4) quadruplex already proposed. For the thermally treated equimolar mixture of d(G(5)T(5)) and d(C(5)A(5)), the fast band with a molecularity of n = 2 corresponds to the Watson-Crick duplex, d(G(5)T(5)). d(A(5)C(5)). The slow band with a molecularity of n = 4 indicates the antiparallel quadruplex [d(G(5)T(5)). d(A(5)C(5))](2), whose observed CD and UV spectra are different from those of [d(G(5)T(5))](4). By electrophoresis, after reannealing the eluted compound [d(G(5)T(5)). d(A(5)C(5))](2), a distinct photograph showing the band splitting of this quadruplex band into the lower duplex and upper quadruplex bands is not possible; but by a transilluminator, we occasionally observe this band splitting with the naked eye. The linear response polarizability tensor calculations for the thus determined structures of the [d(G(5)T(5))](4) quadruplex, the McGavin-like [d(G(5)T(5)). d(A(5)C(5))](2) quadruplex, and the Watson-Crick d(G(5)T(5)). d(A(5)C(5)) duplex are found to qualitatively predict the observed CD and UV spectra.     

Synthesis and luminescence properties of a blue-emitting Sr(3.5) Y(6.5) O(2) (PO(4) )(1.5) (SiO(4) )(4.5) :Eu(2+) phosphor

A novel blue‐emitting Sr_3.5Y_6.5O₂(PO₄)_1.5(SiO₄)_4.5:Eu²⁺ phosphor was synthesized via a solid‐state reaction. Powder X‐ray diffraction (XRD) analysis demonstrated that the Sr_3.5Y_6.5O₂(PO₄)_1.5(SiO₄)_4.5 host had a hexagonal crystal structure in the space group P6₃/m and unit cell parameters a = 9.418 Å, c = 6.900 Å. The as‐prepared phosphor showed a blue emission and all the main emission peaks were located at around 466 nm for different excitation wavelengths of 297, 333 and 391 nm. The temperature dependence of the photoluminescence property was investigated in the range 20–250 °C, and the emission intensity decreased to 71% of the initial value at room temperature on increasing the temperature to 150 °C. According to the classical theory of fluorescent thermal quenching, the activation energy (ΔE) for the thermal quenching luminescence of the as‐prepared Sr_3.45Y_6.5O₂(PO₄)_1.5(SiO₄)_4.5:0.05Eu²⁺ phosphor was determined to be 0.20 eV. Copyright © 2011 John Wiley & Sons, Ltd.     

The frequencies of Gm(1), Gm(2), Gm(4), Gm(12), and Inv(1) in Hessen (Germany)

Summary The serum groups Gm(1) [Gm(a)], Gm(2) [Gm(x)], Gm(4) [Gm(f)]. Gm(12) [Gm(b)] and Inv(1) [Inv(1)] of 2000 sera of healthy blood donors from the land Hesse were examined. The results obtained were compared with those known until now. Three persons, not related to each other, possessed the extremely rare phenotype Gm(-1, 2, 4, 12) [Gm (a-x+b+f+)]. In 0.75% of the cases we found a discordant behaviour of the factors Gm(4) and Gm(12) [Gm(f) and Gm(b)].

---

Zusammenfassung 2000 Seren von gesunden Blutspendern aus Hessen wurden bezüglich der Gamma-Globulin-Serumgruppen Gm(1) [Gm(a)], Gm(2) [Gm(x)], Gm(4) [Gm(f)]. Gm(12) [Gm(b)] und Inv(1) [Inv(1)] untersucht. Die gefundenen Resultate wurden mit den bisher bekannten verglichen. Drei miteinander nicht verwandte Personen wiesen den äußerst seltenen Phänotyp Gm(-1, 2, 4, 12) [Gm(a-x+b+f+)] auf. In 0.75% der Fälle fanden wir ein diskordantes Verhalten der Faktoren Gm(4) und Gm(12) [Gm(f) und Gm(b)].

---

---

Director: Prof. Dr. W. Wachsmuth

---

Director: Prof. Dr. W. Spielmann

---

The nomenclature suggested by WHO at a round-table conference over genes, genotypes and allotypes of immunglobulins is used. The conference took place in Geneva on the 1965 31. 5. to the 5. 6. [5].

---

With technical assistance of S. Mohs.     

Nature and composition of La(III), Ce(III), Pr(III), Nd(III), Sm(III), Gd(III), Dy(III) and Ho(III) complexes of embelin

The isolation and characterization of nine polymeric complexes of the general formula [M(L)_1.5S₂]_n (where M is the metal ion, L the ligand and S the solvent, C₂H₅OH) of La(III) and Ce(III), Pr(III), Nd(III), Sm(III), Gd(III), Tb(III), Dy(III), Ho(III) with.the biologically active compound embelin using elemental and thermal analysis, infrared and electronic spectral studies is reported.