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1.
根据已发表的序列,分别在鸡贫血病毒(CAV)环形基因组DNA(全长2.3kb)的EcoRI位点和BamHI位点的两侧选择适当序列合成两对引物,用PCR技术,从斑点杂交检测到病毒核酸的CAV感染的MDCCRP1细胞基因组DNA中,分别扩增出包含EcoRI和BamHI分割开的病毒基因组两部分(1.5kb和0.8kb)约1.5kb和约1.25kb的两个片段。再将其中相应序列拼接克隆进pUC18载体,获得包含CAV全基因组序列DNA片段的克隆质粒pCAV2.4。酶切分析表明,该质粒具有预期的BamHI位点、PstI位点、HindⅢ位点,而预期的EcoRI位点消失。重组质粒插入DNA片段的两端序列分析表明,质粒pCAV2.4是包含CAV全基因组序列的重组质粒,插入DNA片段序列中的EcoRI位点序列发生了一个碱基突变。 相似文献
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黑曲霉T21是由黑曲霉3.795经诱变育种获得的糖化酶高产菌株,为阐明其高产的分子机制,由黑曲霉3.795克隆了糖化酶结构基因及其5′旁侧序列,并与黑曲霉T21的相应序列进行了比较.由黑曲霉3.795菌丝体分离染色体DNA,Southern杂交分析表明,糖化酶结构基因位于~2.5kb的EcoRⅠ-EcoRⅤ染色体DNA片段上,在此EcoRⅠ位点上游约1.0kb处有一SalⅠ位点.为构建糖化酶结构基因及其5′旁侧序列的基因组文库,该染色体DNA分别用EcoRⅠ+EcoRⅤ和EcoR+SalⅠ消化,琼脂糖凝胶电泳分离并回收长度在1.0kb左右和2.5kb左右的DNA片段,分别与pUC19载体连接后转化入E.coliDH5.用原位杂交方法筛选到了携带糖化酶基因编码区及其1505bp5′旁侧序列的阳性克隆.对克隆片段的DNA序列进行了测定并与黑曲霉T21的相应序列进行了比较,结果表明,在糖化酶基因编码区及其150bp3′非编码区内,未发现碱基差异,但在-340~-1505的5′上游区内发生了9个位置的碱基变化,包括缺失、插入和替换.这些结果表明,黑曲霉T21与3.795的糖化酶产量的差异与其结构基因无关,但可能与其 相似文献
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杂合质粒pIJ8310是链霉菌低拷贝质粒pIJ9221携带了10.8kb来自变铅青链霉菌JT46菌株,编码对噬菌体ΦHAU3抗性的DNA片段。已知Tn4560插到该克隆中3kb的EcoRI或3.3kb的BamHI片段(这两个片段之间有2.5kb的重叠区)上,中断了ΦHAU3抗性基因的表达。已将3.0kb的BcoAI片段分别以两种不同的取向克隆到pBluescriptSK(+)上。核苷酸序列分析揭示出一个1.3kb的ORF的存在,其所编码的478个氨基酸组成的蛋白与λ噬菌体的ea59基因所编码的氨基酸序列显示了高度的同源性,其氨基酸序列的同一性高达37%,相似性达58%,而λ噬菌体的ea59基因是编码赖ATP和单键DNA的核酸内切酶I的,该区域与λ的复制无关。此外,这段具有高度同源性的DNA区域的G+C%只有60%,低于变铅青霉菌DNAG+C%的平均值(74%)。 相似文献
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在克隆了人基因组全长vigilin基因的基础上,对其5‘端转录调控区域再克隆,获得Vig-CG5-7E克隆,再用限制酶Apa1和EcoRⅠ切除3’端约4kb部分,得到Vig-CG5-7E3AE克隆,并对该克隆进行双向测离分析,结果表明:克隆Vig-CG5-7E用ApaⅠ酶消化后,用cDNA上游开始的20个碱基组成 寡聚核苷酸做探针进行分子杂交,呆见一条上于0.1kb杂交带,根据物理图谱分析,位于V 相似文献
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对含有麦迪霉素4″-O-丙酰基转移酶(mpt)基因的BamHI-BamHI8.0kb的DNA片段进行限制性酶切分析,绘制出了含有21个酶切位点的限制性酶切图谱。以含有碳霉素异戊酰基转移酶基因(CarE)的2.4kb DNA片段为探针,经Southern blot分子杂交,将mpt定位于一个EcoRI-EcoRI-PstI3.0kb的DNA片段上,将该片段克隆至大肠杆菌/链霉菌穿梭质粒载体pWHM3 相似文献
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银王亮丝草的试管微繁殖 总被引:2,自引:0,他引:2
1植物名称银王亮丝草(Aglaonema com-mutatumcv."SilverKing”),又名银王粗肋草、银王万年青、银皇帝。2材料类别带腋芽的茎段。3培养条件以MS为基本培养基。芽诱导培养基附加6-BA2.0~2.5mg·L-1(单位下同);芽增殖培养基附加6-BA4.0~4.5及NAA0.01~0.02;生根培养基采用3/4MS附加IBA3.0~3.5及GA33,0。所有培养基蔗糖及琼脂含量分别为3%和0.75%,pH值均调至5.8。培养温度26~28℃,光照8~10h·d-1,光照… 相似文献
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缺氧性肺动脉高压大鼠肺组织中血小板源性生长因子和c—my… 总被引:1,自引:0,他引:1
本研究分析了大鼠肺组织中血小板源性生长因子A链、B链和c-myc原癌基因mRNA。正常肺组织可表达1.7kb的PDGF-AmRNA和3.5kb的PDGF-BmRNA,还有少量2.2kbcmRNA.在缺氧过程中,PDGF-B链mRNA和c-mycmRNA迅速增加,至缺氧14d时,分别为正常的3倍和5倍。而PDGF-AmRNA在缺氧7d时增高,而后又略有降低。结果表明:缺氧的肺组织局生成的PDGF激活 相似文献
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高粱细胞质雄性不育系3197A(3A)在常温条件下是不育的(Figs.11&2),经热激(45℃)诱导不同程度地恢复了育性(Figs.13&4),为研究其不育机理提供了线索。热激2h后,3A中即可产生一类线粒体热激蛋白(HSPs)。其中,分子量为70kD的HSP70含量最高,也最为稳定。不过,3A中HSPs的稳定性弱于保持系3197B(3B)(Fig.2,Panels1~4)。放线菌素D抑制HSPs的合成,而氯霉素无此作用(Fig.2,Panels5&6),表明:HSPs是由核基因编码、在细胞质中合成、再跨膜转运到线粒体中的。3A幼穗经热激后,线粒体的总蛋白量猛增了2.7倍(Fig.3),达到3B的水平,育性亦变为可育的。Fig.4表明:HSP70反义链cDNA(R1)能进入到3B花药细胞中,并与靶RNA(HSC70mRNA)结合,而对照、正义链cDNA(D)链无此反应。由此、再增加一个通用保守序列的反义链cDNA(R2)、共两个探针(R1、R2),可以检测到:3A在常温下没有能力合成HSC70mRNA(Fig.5),而在热激条件下,转变为有能力(Fig.6)。启示:3A在热激条件下由不育转变为可育 相似文献
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用富集文库克隆人胰岛素基因组基因 总被引:1,自引:0,他引:1
通过构建可富集人胰岛素基因的λ噬菌体文库,克隆了人胰岛素基因组基因.首先从中国人血液白细胞中提取到人基因组DNA,用EcoRⅠ和BglⅡ对基因组DNA进行全酶切,经0.4%琼脂糖凝胶电泳,特异回收9.5kb左右的DNA片段.将该片段与λEMBL3/BamHⅠ臂连接,构建成一个特殊的人基因组λ噬菌体文库(富集文库),效价为2×104.同时采用PCR方法及用引物Ⅰ:5′GGACAGGCTACATCAGGAAGAGG3′,引物Ⅱ:5′CTGCGTCTAATTGCAGTAGTTC3′,从人基因组DNA中扩增出一段含胰岛素基因的1.36kbDNA片段,做为放射性标记探针,对文库进行了噬菌斑原位杂交筛选,从1×104个噬菌斑中筛选到一个含人胰岛素基因组基因的阳性克隆,并进一步完成了亚克隆和该基因1732bpDNA序列的测定.结果该基因的1732bpDNA序列包括部分5′端和3′端与国外发表的人胰岛素α型等位基因的序列相同 相似文献
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为研究导致Yunnanese(Aγδβ)0-地贫缺失事件的分子机制,并从3′并入序列中搜寻增强子类序列,使用EMBL3为载体构建了一例缺失杂合子的基因组文库,筛选到来源于异常染色体11并包含Gγ珠蛋白基因区6.7kb序列以及11.5kb3′并入序列(即缺失桥片段)的克隆.此11.5kb序列在正常染色体中位于β珠蛋白基因约下游66~78kb区域.详细分析了这一区域的限制性内切酶图谱.分析了围绕缺失连接区的DNA序列,精确定位缺失的5′端点发生在Aγ珠蛋白基因上游-116~-117碱基之间.确定缺失的3′端点处于一个L1序列内,位于β珠蛋白基因下游~66kb,距离Chinese(Aγδβ)0-地贫缺失3′端点上游~12.2kb的一个EcoRⅠ位点上游413bp.围绕5′和3′端点的序列之间无明显同源性,说明这一缺失代表了体内的非同源重组事件.这一重组事件可能由L1序列介导.缺失3′端点下游序列的克隆分离也为进一步从中搜寻加强子类序列奠定了基础 相似文献
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Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis
It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis. 相似文献
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RICHARD E. NORRIS 《Botanical journal of the Linnean Society. Linnean Society of London》1991,106(1):1-40
Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera. 相似文献
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JOAN VALUÈS MONTSERRAT TORRELL NÚRIA GARCIA JACAS 《Botanical journal of the Linnean Society. Linnean Society of London》2001,137(4):399-407
Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted. 相似文献
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肝癌中HBV和HCV基因和抗原的分布及意义 总被引:1,自引:0,他引:1
采用原位分子杂交方法检测HCV
RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV
RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV
RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV
X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV
C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细 相似文献
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H.-R. GREGORIUS 《Biological journal of the Linnean Society. Linnean Society of London》1984,23(2-3):157-165
For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment. 相似文献
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment. 相似文献