小麦醇溶蛋白mRNA的分离及体外翻译

本文中采用受粉后发育25天的小偃麦种子制备总RNA。然后用高效Oligo(dT)—Celluiose分离Poly(A)—mRNA。根据它在麦胚无细胞蛋白质合成体系中的翻译活性和对体外翻译产物进行聚丙烯酰胺凝胶电泳放射自显影分析说明,我们分离的Poly(A)—mRNA     

光/暗对高粱叶片磷酸烯醇式丙酮酸羧化酶基因表达的调节

高粱幼苗黄化叶片经照光转绿后,其PEP-Case活性提高4～15倍,mRNA含量提高了1.03倍,并测定出PEPCase mRNA的分子量为3.4kb。以等量的总RNA及mRNA进行体外翻译,发现转绿后PEPCase专一性翻译活性提高了51％～53％。这表明光照可以在转录水平上调节PEP-Case的基因表达。     

七星瓢虫卵黄原蛋白基因的表达mRNA的体外转译

七星瓢虫成熟雌虫脂肪体总RNA和poly(A)~+RNA中可转译mRNA的水平约为雄虫和不成熟雌虫的两倍,其中所含的卵黄原蛋白mRNA可在体外转译系统中指导卵黄原蛋白多肽的合成。 雌虫取食人工饲料时,其脂肪体RNA中可转译mRNA的水平很低,不能指导卵黄原蛋白多肽的合成。保幼激素类似物能诱导可转译卵黄原蛋白mRNA的出现。     

鲮鱼垂体Poly(A)~+RNA的分离和纯化及其生物活性鉴定

本文利用快速保温法分离纯化了鲮鱼垂体Poly(A)~+RNA,并首次在北农大“白粒146号”小麦麦胚体系中进行了体外翻译活性测定,对鲮鱼Poly(A)~+RNA其最适镁离子浓度为1.7mmol/L,最适Poly(A)~+RNA浓度为0.12μg/50mL,但钾离子浓度及预保温对蛋白质的合成影响不大。实验结果表明鲮鱼垂体Poly(A)~+RNA的体外蛋白翻译活性达对照组的22倍。由于改进了方法,简化了操作程序,因此使鲮鱼垂体Poly(A)~+RNA的翻译活性大大提高了。     

用钙沉淀法分离植物poly(A)-mRNA

本文对Lendes等人用钙沉淀poly(A)—mRNA的方法作了改进。在不加poly(A)的情况下,适当增加总RNA浓度,用15mM CaCl_2沉淀poly(A)—mRNA。使用改进的方法能够得到大量的poly(A)—mRNA。用改进法制各的poly(A)—mRNA;可以用poly(U)—Sepharose4β或oligo (dT)—Cellulose亲和层析作进一步的纯化,也可以直接作为反转录或翻译的模板.     

拟南芥EBP1蛋白与RNA相互作用的初步研究

基因表达调控是现代分子生物学研究的热点话题,随着研究手段的不断成熟,其RNA转录后调控已成为备受关注的领域。RNA结合蛋白(RBPs)是转录后调控的关键因子,参与RNA可变剪切、RNA稳定、翻译等多个过程的调控。ErbB3绑定蛋白(EBP1)是结构功能高度保守的RNA结合蛋白,可与rRNA、tRNA、mRNA等多种RNA结合,参与调控核糖体生物生成、蛋白质翻译多个过程,但目前已报道的靶RNA甚少,对靶RNA的作用机制目前仍不清楚。通过生物信息学分析发现拟南芥EBP1结合RNA的保守结构域,并在体外可直接结合"GUCUCUCACUGCGACGGCUU"序列;通过RNA免疫共沉淀实验找到了3个靶mRNA(AT1G24792、AT3G25211、AT3G24320);结合核糖体提取及虫草素处理实验发现EBP1可显著调控特定靶mRNA的稳定性及其翻译速率。研究结果不仅证实拟南芥EBP1具有结合RNA的功能,还显示其参与调控靶RNA的转录后事件,为进一步研究EBP1在转录后调控的生物学机制奠定基础。     

家蚕(Bombyx mori)主要血浆蛋白质及其基因表达的研究

本文用SDS-PAGE法观察不同发育阶段蚕血液中主要血浆蛋白质sp、30KP浓度的变化;从不同发育阶段的蚕脂肪体提取RNA和poly(A)~+-RNA,在兔网织红细胞系作体外翻译并检测翻译产物。结果表明,5龄蚕脂肪体mRNA合成蛋白质的速率为初蛹的2倍;5龄及初蛹脂肪体30KP mRNA活性的发育变化与其相应蛋白质在血液中的浓度变化一致;sp-1在5龄幼虫脂肪体内的表达及卵黄原蛋白(Vg)在蚕蛹脂肪体内的表达具有雌特异性,其表达和性特异性大体是在前翻译水平被调节的。     

含尿激酶mRNA的人胚肾培养细胞的poly(A)+RNA的提取及鉴定

人胚肾原代培养细胞经换无血清培养液后4～6天,可产生较高的尿激酶活性,从2ml细胞沉淀中(2×10~5细胞)可提取60～100μgpoly(A)~+RNA。6M尿素酸性琼脂糖凝胶电泳表明,其大小为9S～28S,以此poly(A)~+RNA为模板可用于体外翻译及反转录实验,DNA-RNA点杂交表明,此poly(A)~+RNA含有编码尿激酶的mRNA,通过Northern blot杂交实验,估计mRNA大小在1～3 kb之间,本制品可供应反转录途径获得尿激酶基因之用。     

北京鸭珠蛋白mRNA的分离纯化及其cDNA的合成

 从人工贫血的北京鸭网织红细胞中直接提取总RNA,经Oligo(dT)-纤维素柱层析分离获得珠蛋白mRNA,并经蔗糖密度梯度离心首次得到了电泳单一条带的北京鸭球蛋白mRNA。从凝胶电泳以及蔗糖密度梯度离心鉴定其沉降系数为9S。在麦胚无细胞体外翻译体系中测定了它们的蛋白翻译活力。鸭珠蛋白mRNA促进了~3H-亮氨酸参入新生蛋白的活力,达到对照组的10倍。所翻译的蛋白产物在SDS-聚丙烯酰胺凝胶上的电泳行为与天然鸭珠蛋白一致。 经Oligo(dT)-纤维素及蔗糖密度梯度离心提纯的珠蛋白mRNA,在AMV反转录酶及DNA聚合酶的作用下,分别合成了单链及双链cDNA。其双链链长,经凝胶电泳分析,约为500碱基对。     

用低浓度硫氰酸胍提取高质量植物RNA

目前分离RNA的方法很多, 但大多是基于动物材料建立的方法, 而针对植物材料的方法并不多见. 建立了一种从植物材料中分离高质量RNA的硫氰酸胍/氯化锂/热酚法. 与其他硫氰酸胍的方法相比, 该方法所用硫氰酸胍的浓度仅为现有方法的1/40 (0.1 mol/L),降低了实验成本, 且所得RNA质量令人满意. 用该方法分离的RNA在琼脂糖凝胶电泳上可清晰分出4条核糖体RNA (rRNA)带; 用此RNA进行RNA印迹或分离mRNA进行体外翻译试验, 均获得很好效果.     

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.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

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.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

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.     

肝癌中HBV和HCV基因和抗原的分布及意义

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     

Selection with two alleles and complete dominance

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.