基于网络层次分析法的酵母转化实验策略优化

结合建立的酵母转化实验影响指标体系,利用网络层次分析法(ANP)解决了酵母转化实验策略优化的问题．通过对已有成果进行研究总结,结合前期基础实验以及 对该领域专家学者的咨询意见,建立了酵母转化实验影响指标体系,并基于管理学的ANP理论对实验方法进行了两两比较,分析得出电击穿孔为最优酵母转化方法．实验表明,利用脂质体法得到的转化子为31个/μg质粒DNA,利用电击穿孔法在电压为2.0 kV、1.5 kV时得到的转化子分别为37、29个/μg 质粒DNA,说明电击穿孔实验转化率相对较高,且电压的改变对转化率影响较大,与指标体系分析结果相符．故该指标体系与分析方法可为毕赤酵母转化实验策略优化提供有效的理论依据,其思路、体系、理论和方法可为同类实验所借鉴．     

电击法锈导的欧白英原生质体转化和转化植株再生

本研究使用电击法成功地将带有标记基因ＮＰＴⅡ的质粒ｐＣａＭＶＮＥＯ转入欧白英的原生质体,并获得了再生转化植株。通过用ｐＤＷ２质粒进行的ＣＡＴ基因短暂表达研究,确定了欧白英原生质体转化的电击条件为：电容３０ｎＦ、电场强度１５００Ｖ／ｃｍ、时间衰变常数５９．４微秒;质粒ＤＮＡ浓度为２０μｇ／２×１０＾６原生质体。在以上条件下,欧白英原生质体的相对转化率为１２．４％,绝对转化率为２．４×１０－４。在大多     

蓝藻的一种人工转化系统研究

本研究用溶菌酶处理ＳｙｎｅｃｈｏｃｏｃｃｕｓＰＣＣ６３０１细胞诱导人工感受态,并用外源质粒ｐＢＲ３２５转化受体细胞表达氯霉素抗性,转化频率接近５×１０￣（－５）转化子／细胞。用转化子ＤＮＡ再进行次级转化,转化频率可达５×１０￣（－４）转化子／细胞。这比以前的研究者对同种藻株,用克隆的ＤＮＡ、通过生理感受态进行转化得到的转化频率要高。ＤＮＡ电泳、次级转化和斑点杂交证明外源质粒通过单交换已经整合到了受体细胞染色体上。这些结果表明,人工转化系统是有效的,并具有可重复性。对于影响转化的一些条件,如ＤＮＡ与细胞保温时间、藻龄、光照或黑暗培养,也同时进行了研究。     

外源质粒电击转入Pseudomonas putida的条件研究

以自行筛选的恶臭假单胞菌（Pseudomonas putida）(命名为Rs198,Genbank登录号为FJ788425）为受体菌,将具有卡那霉素抗性标记的大肠杆菌假单胞菌穿梭质粒PDSK519通过电转化法导入到受体菌中,对细胞生长状态、电转化温度、质粒DNA及感受态细胞浓度、电击电压及电转化介质给予转化效率的影响进行研究。结果表明,在细胞生长至OD600为0.5左右时收集菌体,在低温条件下制备浓度为 4.6×1012/ml 的感受态细胞,以0.3mol/L的蔗糖为电转化介质,在13kV/cm的场强下电击能获得较高的转化效率,最高可达1.3×107个转化子/μ g DNA。为构建恶臭假单胞的遗传转化系统,利用基因工程手段为该菌的进一步研究奠定了理论基础。     

植病生防菌成团泛菌电击转化条件的研究*

研究了植病生防菌成团泛菌（Ｐａｎｔｏｅａ ａｇｇｌｏｍｅｒａｎｓ）的电击转化条件。结果表明受体菌固体培养、生长到对数早期时的细胞转化效果最好;质粒ＤＮＡ分子量增加１０培,转化效率降低１００倍;质粒ＤＮＡ浓度在０．０１－１μｇ／μＬ范围内其变化与转化频率呈正比,与转化效率成反比;从成团泛菌中提纯的质粒ＤＮＡ比从大肠杆菌中提纯的相同质粒ＤＮＡ转化成团泛菌的效率高出３０倍;最佳电击条件为场强１５ｋＶ／ｃ     

具有真细菌基因启动子活性的盐生盐杆菌质粒DNA片段

利用大肠杆菌启动子探测质粒ｐＫＫ２３２－８为载体、用两组限制性内切酶ＢａｍＨＩ－ＳａｌＩ和ＨｉｎｄⅢ－ＳａｌＩ分别消化盐生盐杆菌Ｊ７（Ｈａｌｏｂａｃｔｅｒｉｕｍｈａｌｏｂｉｕｍ）的质粒ｐＨＨ２０５,在体外进行重组,转化Ｅ．ｃｏｌｉＨＢ１０１感受态细胞,在含氨苄青霉素和氯霉素的选择平板上筛选转化子,并从随机挑选的２０株转化子中,获得抗氯霉素水平达到１１０μｇ／ｍｌ的转化子Ｔ１和Ｔ２,所含重组质粒分别被命名为ｐＪＨ和ｐＪＢ。经限制性酶切分析及杂交分析表明,ｐＪＨ质粒上插入了一段来源于ｐＨＨ２０５质粒的ＤＮＡ片段,其大小为８００ｂｐ左右。通过重新转化实验进一步表明,该ＤＮＡ片段在大肠杆菌中具有启动子功能,从而证明,在古细菌（盐生盐杆菌）的质粒ＤＮＡ中存在具有真细菌（大肠杆菌）基因启动子活性的ＤＮＡ片段。     

用电注入质粒DNA转化完整酵母细胞的新方法

已经发现,用电场脉冲可以将质粒DNA(YEp13)引入完整的酵母细胞。电注入法的转化频率决定于起始电场强度、放电电容器的电容及应用的脉冲。用起始电场强度5kv/cm和电容/1μF的3个连续脉冲得到转化子数最多(90±20个/μgDNA)。电注入方法简单,且在转化处理后2-3天内即可得到转化子。     

niaD基因与uidA基因共转化糖化酶高产菌株Aspergillus nigerT21

从Ａｓｐｅｒｇｉｌｌｕｓ ｎｉｇｅｒ Ｔ２１分离到自发性的氯酸盐抗性株,再经氮源生长试验获得硝酸盐还原酶缺陷的ｎｉａＤ突变体Ｎ４４。用含有ｎｉａＤ的质粒ｐＳＴＡ１０转化Ｎ４４,转化频率为５个／μｇ（转化子／ＤＮＡ）。转化子的Ｓｏｕｔｈｅｒｎ印迹分析表明ｎｉａＤ基因同源整合到Ｎ４４的染色体ＤＮＡ中。ｐＳＴＡ１０与含葡萄苷酸酶基因（ｕｉｄＡ）的质粒ｐＮＯＭ１０２共转化Ｎ４４,共转化频率为４０％。共转化     

小麦G—型细胞质雄性不育的育性恢复因子Rf3的研究Ⅱ.总体DNA构建提莫菲维小麦基因组文库

分别用ＰｓｔⅠ、ＨｉｎｄⅢ和ＥｃｏＲⅠ三种限制性内切核酸酶切割提莫菲维小麦基因组ＤＮＡ,再与用相应酶处理的载体ｐＵＣ１１９连接,然后转化到大肠杆菌菌系ＤＨ５α之中。转化效率为１．２×１０４／μｇ。经α互补检测,初步筛选重组子。随机挑取单菌落分别培养,提取质粒,经琼脂糖凝胶电泳和斑点杂交,最后确定重组子,重组率为４３．８％。粗略地估计了克隆片段在基因组中的拷贝数     

生米卡链霉菌丙酰化酶基因定位及其核苷酸序列测定

我们已往的工作已经确定在重组质粒ｐＩＪＭ９中,生米卡链霉菌来源的４．１６ｋｂ插入ＤＮＡ片段带有丙酰化酶基因,为了进一步确定该基因在４．１６ｋｂ插入片段中的位置,我们以ＢａｍＨＩ对ｐＩＪＭ９进行酶切,在此基础上进行缺失重组亚克隆,在所获得的转化子中,Ｎｏ．５转化子含重组质粒ｐＩＪＭ９５,分子量为５．０ｋｂ,插入ＤＮＡ片段为０．５３ｋｂ,以Ｎｏ．５转化子对螺旋霉素进行生物转化实验,其转化产物经薄层层析     

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.