Purification of immunoglobulins G by protein A/G affinity membrane chromatography

An affinity membrane grafted with protein A/G or protein A was characterized for human and mouse immunoglobulins G purification. Breakthrough curves up to ligand saturation were measured and used to study the effects of flow velocities, feed solution concentrations and protein A/G versus protein A membranes. Increased flow-rate did not decrease the amount of IgG bound to the membranes. Increased feed solution concentration allowed more IgG to bind prior to breakthrough. Kinetic parameters for immunoglobulins G sorption to immobilized protein A were measured in batch experiments. The static binding capacity was determined to be 6.6 mg ml⁻¹ membrane volume. Finally, this affinity membrane was used to purify IgG from cell culture supernatant. The electrophoresis of the purified IgG fractions did not show any contaminant.     

Sequence specificity of inter- and intramolecular G-quadruplex formation by human telomeric DNA

Human telomeric DNA consists of tandem repeats of the sequence 5'-d(TTAGGG)-3'. Guanine-rich DNA, such as that seen at telomeres, forms G-quadruplex secondary structures. Alternative forms of G-quadruplex structures can have differential effects on activities involved in telomere maintenance. With this in mind, we analyzed the effect of sequence and length of human telomeric DNA on G-quadruplex structures by native polyacrylamide gel electrophoresis and circular dichroism. Telomeric oligonucleotides shorter than four, 5'-d(TTAGGG)-3' repeats formed intermolecular G-quadruplexes. However, longer telomeric repeats formed intramolecular structures. Altering the 5'-d(TTAGGG)-3' to 5'-d(TTAGAG)-3' in any one of the repeats of 5'-d(TTAGGG)(4)-3' converted an intramolecular structure to intermolecular G-quadruplexes with varying degrees of parallel or anti-parallel-stranded character, depending on the length of incubation time and DNA sequence. These structures were most abundant in K(+)-containing buffers. Higher-order structures that exhibited ladders on polyacrylamide gels were observed only for oligonucleotides with the first telomeric repeat altered. Altering the sequence of 5'-d(TTAGGG)(8)-3' did not result in the substantial formation of intermolecular structures even when the oligonucleotide lacked four consecutive telomeric repeats. However, many of these intramolecular structures shared common features with intermolecular structures formed by the shorter oligonucleotides. The wide variability in structure formed by human telomeric sequence suggests that telomeric DNA structure can be easily modulated by proteins, oxidative damage, or point mutations resulting in conversion from one form of G-quadruplex to another.     

3种甘草属植物的核型研究

用光学显微镜观察了豆科（Leguminosae）甘草属（Glycyrrhiza）3种荒漠植物的染色体,研究结果表明,3种植物的体细胞染色体数目2n=16,核型公式分别为：刺果甘草（G.pallidiflara）k(2n)=2x=16=4M+8m+4sm,光果甘草（G.glabra）k(2n)=2x=16=10M+6m,胀果某草（G.inflata）k(2n)=2x=16=6M+6m+4sm。     

刺槐对3种抗生素敏感性的测定

匈牙利刺槐对卡那霉素不敏感,对G418敏感,而对潮霉素极为敏感.如以NPTII基因为刺槐基因工程的选择性标记基因,则G418是理想的抗生素抗性选择试剂;如用HPT基因,则潮霉素是合适的抗生素抗性选择试剂.     

鳞盖肉齿菌的抗肿瘤细胞增殖活性及抗肿瘤细胞增殖机制的研究

为了从鳞盖肉齿菌(Sarcodon scabrosuskarst)的二氯甲烷提取物中分离纯化得到Sarcodonin G,对Sarc-odonin G进行抗肿瘤细胞增殖活性及其抗肿瘤细胞增殖机制的研究。本文利用MTT实验法测定Sarcodonin G对HeLa细胞株增殖的抑制率,检测其抗肿瘤细胞增殖的效果;利用流式细胞术检测Sarcodonin G对HeLa细胞的凋亡的影响;利用电镜技术观察Sarcodonin G对HeLa细胞形态学改变。结果发现,Sarcodonin G对体外培养的HeLa细胞的增殖具有明显抑制作用,其IC50为7.19μmol/L,并有较好的剂量依赖关系;流式细胞学检查发现Sarcodonin G处理后的HeLa细胞出现凋亡现象;超微结构发现Sarcodonin G处理后的HeLa细胞的胞核染色质浓缩,边集,核固缩及形成新月体,线粒体肿胀,空泡样变,胞浆出现大量空泡等凋亡的形态学改变。结果表明,鳞盖肉齿菌的纯化物Sarcodonin G能抑制体外培养的HeLa细胞的增殖,并初步推测Sarcodonin G有可能是通过诱导HeLa细胞的凋亡来实现其抗肿瘤增殖作用的。     

中国鹿花菌属志略

近年来,国外许多学者对鹿花菌属 Gyromitra Fr.的一系列研究导致该属传统的概念、范围发生了很大的变动。本文在接受了 Harmaja 将 Discina 和 Neogyromitra 合并在Gyromitra 名下的观点的基础上,报道了近年来作者在我国鹿花菌属研究中所确认的8个种。其中乳白鹿花菌 Gyromitra lactea 和新疆鹿花菌G.xinjiangensis 为新种,含糊鹿花菌G.ambigua、帚状鹿花菌 G.fastigiata 和亮鹿花菌 G.splendida 为我国首次报道。我国文献中所记载的鹿花菌 G.esculenta 在我国是否存在,尚需进一步调查。     

玉米粉在赤芝液体培养中的应用研究

以菌丝含量为主要指标,研究了6种营养物质对赤芝深层培养的影响,试验结果表明,赤芝500ml小规模发酵最适培养基配方为:玉米粉4.0%,糖蜜1.5%,黄豆粉2.0%,酵母膏0.2%,KH2PO40.2%,MgSO4.7H2O 0.15%,pH 6.0。接种量为10%时在150r/min摇床上26℃振荡培养5d,菌丝含量最高,为25.1g/L;玉米粉是赤芝生长的主要因素,糖蜜是次要因素,玉米粉可以作为赤芝发酵培养基的主要原料。     

灵芝栽培培养基筛选试验

以灵芝(Ganoderma lucidum)之赤芝(Ganoderma lucidum Karst)、韩芝(Ganoderma luci- dum H.G)为试验品种,培养基以棉壳98%、石膏1%、蔗糖1%为对照,另外以棉壳、麸皮、木屑、氨基酸不同含量配制4种培养基作比较栽培试验。结果表明:2号培养基出菇产量高、菇形好,对赤芝、韩芝都适宜;赤芝、韩芝对培养基的要求略有差异,韩芝在含麸皮量高的基质上栽培更为适合。     

异种化人肾细胞癌特异性抗原G250真核表达载体的构建及表达

目的：构建含有人肾细胞癌特异性抗原G250（CAⅨ）主要T细胞表位区域、猴和鼠CAⅨ部分片段区域融合基因tG250的真核表达质粒,并在猴肾COS7细胞中表达。方法：通过基因合成和PCR技术构建人、猴和鼠G250区域融合基因tG250,将其插入含有人Igκ链前导信号肽（sig）、人IgG-Fc和糖基磷脂酰肌醇（GPI）锚定信号肽融合基因序列的细胞膜锚定修饰真核表达载体pCI-Fc-GPI中,继而又将酶切后的sig-tG250-Fc-GPI融合基因导入含有细小病毒内部核糖体结合位点（IRES）基因且可以共表达人GM-CSF和B7.1融合基因的真核表达载体pVAX1-IRES-GM/B7中;将构建的重组质粒pVAX1-sig-tG250-Fc-GPI-GM/B7转染COS7细胞,利用流式细胞仪和免疫荧光检测其表达。结果：测序结果表明tG250融合基因序列正确,PCR和酶切鉴定证明已将其连入真核表达载体pVAX1-IRES-GM/B7中;流式细胞仪和免疫荧光的检测结果显示,重组质粒pVAX1-sig-tG250-Fc-GPI-GM/B7在COS7细胞中得到很好的表达。结论：构建了重组质粒pVAX1-sig-t G250-Fc-GPI-GM/B7,且在COS7细胞中有效表达,为以G250为靶点的抗肾细胞癌基因疫苗的构建与功能研究奠定了基础。     

Screening of mitochondrial mutations in Saudi women diagnosed with gestational diabetes mellitus: A non-replicative case-control study

IntroductionAmong metabolic disorders, gestational diabetes mellitus (GDM) is specified as hyperglycemia caused by glucose or carbohydrate intolerance defects. GDM is distinguished by oxidative stress, and has been connected to mitochondrial dysfunction. Previous studies have documented the relation between A12026G, A8344G and A3243G mutations in ND4, tRNA^Leu(UUR), and tRNA^Lys genes in different modes of diabetes.AimThe purpose of this study was to investigate into the relationship between GDM women and common mitochondrial mutations including A12026, A8344G, and A3243G in Saudi women.MethodsIn this case-control study, we have opted 96 GDM and 102 non-GDM pregnant women and DNA was extracted using EDTA blood and based on specific primers, Polymerase Chain Reaction was followed and then Restriction Fragment Length Polymorphism (RFLP) analysis was performed. Restriction enzymes was cross-checked with Lambda DNA and 10% of the purified PCR products were performed the Sanger sequencing analysis to reconfirm the RFLP analysis of the studied results.ResultsNone of the heterozygous and homozygous mutations were not observed in our study. All the subjects were turned to be homozygous normal genotypes.ConclusionThis study confirms that A12026, A8344G, and A3243G mutations have no role in the Saudi women with GDM.     

Inhibition of apoptosis in serum starved porcine embryonic fibroblasts

In nuclear transplantation, serum starvation is a general method to synchronize donor cells at the quiescent stage (G(0)) of the cell cycle. However, serum starvation during culture of mammalian cells may induce cell death, especially through apoptosis, thus contributing to the low efficiency of nuclear transplantation. This study was performed to characterize apoptosis during serum starvation and to determine the effects of apoptosis inhibitors such as a protease inhibitor [alpha(2)-macroglobulin (MAC)] and antioxidants [N-acetylcysteine (NAC), glutathione (GSH)] on serum starved porcine embryonic fibroblasts (PEF). PEF, collected from day 25-30 porcine fetuses, were cultured for 5 days in media containing 0.5% FBS to induce quiescence. Serum starved PEF showed typical morphology of apoptotic cells and stained for DNA fragmentation by TUNEL assay (26.7%). All apoptosis inhibitors tested in this study significantly (P < 0.05) reduced apoptosis of serum starved PEF, with antioxidants having better results (MAC: 7.4% vs. NAC: 1.0%, and GSH: 0.8%). Equally and importantly, the treatment with apoptosis inhibitors did not change the proportion of G(0)/G(1) stage cells. Therefore, the addition of MAC and antioxidants during serum starvation of PEF reduces apoptosis of quiescent fibroblasts and may contribute to increasing the efficiency of nuclear transplantation by improving the quality of donor nuclei.     

Comparative study on the infection rates of different laboratory strains of Glossina species by Trypanosoma congolense

Teneral Glossina morsitans centralis Machado, G.austeni Newstead, G.palpalis palpalis Robineau-Desvoidy, G.p.gambiensis Vanderplank, G.fuscipes fuscipes Newstead, G.tachinoides Westwood and G.brevipalpis Newstead, from laboratory-bred colonies, were fed at the same time on the flanks of ten goats infected with Trypanosoma congolense Broden isolated in Tanzania or in Nigeria. The seven tsetse species were infected over the range 0.3-49.2%. Survival of both T.congolense isolates was best in G.m.centralis, poorest in G.austeni and the four palpalis group tsetse, with G.brevipalpis intermediate. It is suggested that there are differences in the gut of different laboratory-bred cultures of Glossina Westwood species and subspecies such that T.congolense parasites can survive better in the gut of some than in others and undergo cyclical development to metacyclics in the hypopharynx.     

植物G蛋白偶联受体及其在信号转导中的作用

G蛋白偶联受体(G protein-coupled receptors,GPCRs)是具有7个跨膜螺旋的蛋白质受体,是人体内最大的蛋白质超家族.GPCRs能调控细胞周期,参与多种植物信号通路以及影响一系列的代谢和分化活动.简要介绍了GPCR和G蛋白介导的信号转导机制,GPCRs的结构和植物GPCR及其在植物跨膜信号转导中的作用,并对GPCR的信号转导机制及植物抗病反应分子机制的研究提出展望.     

Two New Species of Gypsophila L. (Caryophyllaceae) from China

Gypsophila huashanensis Y. W. Tsui et D. Q. Lu and G. spinosa D. Q. Lu (Caryophyllaceae) are described as new from China.     

中国灵芝科的分类研究 Ⅺ.灵芝亚属灵芝组

灵芝(Ganoderma lucidum)远在东汉(纪元102—200)年间的《神农本草经》中就有记载。中国劳动人民认识它远在此之前。国外最早的记载是1781年W.Curtis的“Boletus lucidus W.Curt.,t.no.224”.此种模式标本已不存在。Karsten(1881)在赫尔辛基植物博物馆留下一号标本作为新模式(neoty pc)。不同的分类学家认为在同一种不同的标本上皮壳构造可能有变化,甚致在同一号标本中也有变化。菌肉的颜色也有变化。或许菌肉颜色变暗是由于纬度和海拔高度的不同,平均温度渐渐增加所致(steyaert,1972)。有些分类学家认为菌柄的有无不能作分类的依据。Steyaert(1975)证明甚致硬皮灵芝(Ganoderma tornatum coinplex)的孢子因纬度和海拔高度的不同有改变。总而言之,我们可以说灵芝复合种(Ganoderma complex)在自然界的存在是不稳定的。诚然,这是一条自然规律。在自然界每一个生物种都是连续不断的变化着。但也相对的稳定。它们在自然界的矛盾中生长和发育。本文作者同意其他分类学家的意见,即在灵芝属中孢子诸性状是唯一地分类根据。种与种之间孢子的性状是清楚的不同。同时他建议应用每一个种的综合性状作分类依据或许是正确的。因为不同的分类学家对灵芝有人持广义的概念,也有人持狭义的概念。对它的亲缘种自然也有不同。下列种类现作者认为可能是它的亲缘种类。它们是:无柄灵芝;弱光泽灵芝;四川灵芝;紫芝以及松杉树芝。现作者假定灵芝在本组中是一个中心种。其它亲缘种围绕它而诞生。于是形成一个较大的种群。也有分类学家称之为灵芝复合种。长期以来灵芝复合种已经成为分类上的困难问题。有人认为成了本组分类的绊脚石。作者研究本组真菌已有多年。根据他的经验认为持种的狭义概念大概更合乎自然规律。对于以上观点作者拟在另文中阐述。本文因限于篇幅,只报道4个新种。其中1新种应隶属紫芝组。它们是:大青山灵芝(Ganoderma daiqingshanense Zhao),昆明灵芝(G.kunmingense Zhao),多分枝灵芝(G.ramosissimum),澄海灵芝(G.chenghaiense Zhao)。以上所有引证标本都保藏于中国科学院微生物研究所真菌标本室。     

海南省裸伞属新种和中国新记录

本文报道了海南省裸伞属(Gymnopilus Karsten)的1个新种和3个中国新记录种。该新种为海南裸伞(Gymnopilus hainanensis T. H. Li et W. M. Zhang)。新记录种为栎裸伞(Gymnopilus dryophilus Murrill),厄尔裸伞(G. earlei Murrill)和细鳞裸伞(G. parvisquamulosus Hesler)。所有标本保藏于广东省微生物研究所真菌标本室(HMIGD)内。     

艾滋病患者APOBEC3G 基因的克隆和真核表达

目的 建立艾滋病( AIDS) 患者载脂蛋白B mRNA 编辑酶催化多肽样蛋白3G( APOBEC3G) 的真核表达体系。方法 采用反转录-聚合酶链反应( RT-PCR) 技术从AIDS 患者外周血单个核细胞( PBMC) 中获取APOBEC3G 基因编码区, 将其克隆到pMD18-T载体上, 测序验证正确后再将其转接入真核表达载体pEGFP-N1 中, 然后将重组质粒pEGFP-N1-A3G 转染HEK293T细胞, 分别用RT-PCR 法和蛋白印迹法( Western 印迹法) 验证APOBEC3G 在mRNA 和蛋白水平的表达。结果 从AIDS 患者体内克隆的APOBEC3G 基因编码区长度为1 154 bp, 测序结果与GenBank 中APOBEC3G 参考序列( NM021822) 比对发现存在2 处差异, 分别位于mRNA 第588 位和746 位碱基处。重组质粒pEGFP-N1-A3G转染HEK293T 细胞, 在荧光显微镜下观察到融合蛋白A3G-EGFP的表达, RT-PCR 法和Western blot 法分别验证了蛋白在mRNA 和蛋白水平的表达。结论 成功构建了AIDS 患者APOBEC3G 蛋白的真核表达体系, 为进一步研究APOBEC3G 在HIV-1 感染中的作用奠定了基础。     

AM真菌在煤矿废弃物中生态适应性的初步研究

以粉煤灰、草炭、蛭石、河沙为培养基质,分别对4种不同AM真菌:Glomus mosseae,G.diaphanum,G.intraradices和G.versiforme的生态适应性进行研究。结果表明,菌根的侵染率、孢子密度和菌丝长度分别与菌根真菌种类、培养基质状况及寄主植物种类有关。4种基质的扩繁效果顺序为:河沙>粉煤灰>草炭>蛭石。G.mosseae和G.diaphanum在基质中的产孢量和菌丝长度优于G.intraradices和G.versiforme,可作为优势菌株。4种菌根真菌在粉煤灰中对寄主的侵染率均达到60%以上,粉煤灰作为菌根真菌培养基质具有更大潜力和实际应用价值。     

Replication of bacteriophage f1: A complex containing gene II protein in gene V mutant-infected bacteria

A dense complex has been isolated from bacteria infected with gene V amber mutant f 1 bacteriophage. The major protein in this complex is the f 1 bacteriophage-specific gene II protein. Other proteins in the complex include the f 1 bacteriophage coat protein and proteins which migrate on sodium dodecyl sulfate/polyacrylamide gel electrophoresis with the f1 bacteriophage-specific gene III, gene IV and X protein. A protein of approximately 20,000 M_r is also present in the complex. Examination of bacteria infected with gene V mutant f1 bacteriophage revealed the complex as a densely staining amorphous body which appears to be associated with the cytoplasmic membrane. Bacteria infected with f1 bacteriophage that contain amber mutations in genes other than gene V do not contain this complex.