甘油对杜氏盐藻电击转化的影响

通过在HEPES电击缓冲液中添加不同浓度的甘油,讨论了甘油对电转前细胞存活率的影响;通过在盐藻培养基中添加不同浓度的甘油,讨论了甘油对盐藻细胞生长的影响;使用含有不同浓度甘油的HEPES缓冲液介导质粒载体转入盐藻细胞,比较了甘油对于转化率的影响。实验结果表明,在电击缓冲液中添加0.5mol/L甘油能有效提高细胞存活率,促进转化细胞恢复生长,从而获得最佳转化效果。因此,0.5mol/L甘油可作为杜氏盐藻电击转化过程中一种良好的稳渗剂。     

杜氏盐藻玻璃珠新型转化方法的建立

首次采用玻璃珠法成功转化了杜氏盐藻(以下简称盐藻),转化细胞经染色后呈现蓝色,表明外源报告基因GUS得到了成功的表达。同时还进行了转化时间、转速、PEG和质粒DNA浓度等因素对转化影响的分析,优化了转化条件。结果显示最佳的转化条件为:在800μL盐藻(106个细胞/mL)中加入150μLPEG和90μL质粒,在300mg玻璃珠存在的条件下,于转速2400r/min涡旋12s能够得到较为理想的转化结果。该方法与已报道的转化方法相比,具有操作简便、省时快捷、不需要昂贵消耗试剂和仪器设备、比较经济等优点。此方法的建立为深入研究盐藻基因工程提供了有力的工具。     

高渗提高凝结芽孢杆菌P4-102B菌株的电击转化效率

【目的】凝结芽孢杆菌(Bacillus coagulans)是一种非常有工业应用前景的微生物,但遗传转化的困难是限制对其进行代谢工程改造的关键因素。本研究主要考察生长培养基、电击缓冲液、复苏培养基中添加高渗剂如山梨醇、甘露醇等对凝结芽孢杆菌转化效率稳定性的影响,并对凝结芽孢杆菌电击转化条件进行优化。【方法】用穿梭质粒p NW33N电转化凝结芽孢杆菌P4-102B,系统地考察高渗条件下细胞生长阶段、感受态菌体浓度、电击缓冲液组分和复苏培养基组成等因素对转化效率的影响。【结果】同一电场压力下高渗体系转化效率较低渗体系明显提高且稳定性较好,菌体在含有0.5 mol/L山梨醇的LB培养基中生长到OD600为0.8时收集,用SMG[0.5 mol/L山梨醇,0.5 mol/L甘露醇,10%(质量体积比)甘油]电击缓冲液洗涤菌体4次制备感受态,在固定电场强度14 k V/cm、脉冲时间5 ms、1 mm规格的电转杯进行电击转化,电转化后立即加入含有0.5 mol/L山梨醇和0.38 mol/L甘露醇的LB复苏培养基培养,能够获得最佳的转化效率2.7×102 CFU/μg DNA。【结论】使用高渗电击转化法能够提高电击转化的稳定性和重复性,并且可以获得较高的转化效率。     

外源ble基因在杜氏盐藻中的瞬时表达

利用电击法将带有ble基因的pSP124S转入杜氏盐藻细胞内进行瞬时表达,研究了外源基因在盐藻内的存留及表达情况,确定了合适的电击转化条件,发现利用电击法可以使大量的质粒导入盐藻细胞,质粒在细胞中逐渐降解但至少96h内可以检测得到,外源启动子能够使ble基因有效转录,转录至少可以持续72h,ble基因能够在盐藻细胞中正确翻译,可以作为盐藻遗传转化研究的筛选标记。     

外源ble基因在杜氏盐藻中的瞬时表达

利用电击法将带有ble基因的pSP124S转入杜氏盐藻细胞内进行瞬时表达．研究了外源基因在盐藻内的存留及表达情况,确定了合适的电击转化条件,发现利用电击法可以使大量的质粒导入盐藻细胞,质粒在细胞中逐渐降解但至少96h内可以检测得到。外源启动子能够使ble基因有效转录,转录至少可以持续72h,ble基因能够在盐藻细胞中正确翻译,可以作为盐藻遗传转化研究的筛选标记。     

丁草胺对杜氏盐藻生理生化的影响

采用室内模拟培养试验方法,研究了酰胺类除草剂丁草胺对杜氏盐藻(Dunaliella salina)生长和生理生化的影响。结果表明,低浓度的丁草胺对杜氏盐藻生长速率有促进作用,而高浓度的丁草胺对杜氏盐藻生长速率有显著的抑制作用,且随着浓度的加大,杜氏盐藻生长速率逐渐降低。丁草胺对杜氏盐藻藻细胞的光合色素含量、可溶性蛋白、超氧化物歧化酶(SOD)和过氧化物酶(POD)活性等也有影响。     

高浓度钾抑制杜氏盐藻生长的生理机制

在含１ｍｏｌ／ＬＮａＣｌ的杜氏盐藻（Ｄｕｎａｌｉｅｌａｓａｌｉｎａ（Ｄｕｎａｌ）Ｔｅｏｄ．）培养液中加入５０ｍｍｏｌ／Ｌ以上的ＫＣｌ可观察到Ｋ＋对杜氏盐藻生长有明显的抑制作用,而当ＫＣｌ达１００ｍｍｏｌ／Ｌ时,杜氏盐藻的生长被完全抑制。另一方面,当培养液中缺乏Ｋ＋时,杜氏盐藻的生长也被显著抑制。在正常培养条件下,伴随着杜氏盐藻的生长,培养液的ｐＨ由８左右升高至１０左右,而高浓度Ｋ＋则显著抑制杜氏盐藻培养液ｐＨ的升高;而在培养液ｐＨ为７．０至１０．０的范围内,不同ｐＨ对杜氏盐藻的生长无明显影响。将杜氏盐藻在高浓度Ｋ＋条件下预处理１２ｈ以上,杜氏盐藻的光合放氧速率显著下降,光合速率下降的程度与Ｋ＋浓度的高低和预处理的时间长短呈正相关。高浓度Ｋ＋处理也引起杜氏盐藻叶绿素含量的显著下降。对经高浓度Ｋ＋预处理的杜氏盐藻的光合放氧速率与培养液中ｐＨ变化同时进行测定的结果表明,Ｋ＋抑制杜氏盐藻光合速率的同时也显著抑制了光照条件引起的培养液ｐＨ的上升。实验结果表明,Ｋ＋抑制杜氏盐藻光合作用以及抑制杜氏盐藻生长与Ｋ＋影响跨盐藻质膜的质子运输之间可能存在一定关系。     

杜氏盐藻和亚心形扁藻在不同接种密度和氮浓度下的细胞群体生长

在氮浓度为1.4、14、和140 mg·L^-1下,对杜氏盐藻(Dunaliella salina)和亚心形扁藻(Platymonas subcordiformis)进行细胞接种比例为10:0、7:3、5:5、3:7和0:10的培养试验,研究不同海洋经济微藻种间混合培养的细胞群体生长效应。结果表明,杜氏盐藻与亚心形扁藻细胞群体的生长随着氮浓度的增加而提高;两种藻在氮浓度分别为14和140mg·L^-1时混合培养的细胞群体生长明显优于单独培养。中、高氮下杜氏盐藻与亚心形扁藻以7:3的接种比例混合时,微藻的细胞群体生物量和胞内物质含量相比单独培养和其他比例均有显著提高,如叶绿素a含量比单独培养分别提高1.17倍和7.77倍;蛋白质含量比单独培养分别提高19.1%和195.3%等。而低氮浓度下,藻的生长受到氮浓度的限制。     

变形链球菌电击转化条件的优化研究

目的为增强对变形链球菌的电击转化效率,探索常用的胞壁弱化剂甘氨酸在电击转化中的加入模式。方法以氨苄青霉素抗性的pGL3 basic质粒作为外源DNA,通过电击转化导入变形链球菌参考株UA159内,并在选择性培养基上筛选阳性转化克隆,以优化筛选出最佳的甘氨酸加入浓度与模式,同时比较了不同电击方案转化效率的差异。结果在变形链球菌对数生长期后加入终浓度为10%的甘氨酸可有效地增强电击转化的效率;而不同的电转电压的选择对于转化效率的影响,在本实验中差异未见显著性。结论研究证实了甘氨酸作为胞壁弱化剂可增强对变形链球菌的转化效率,并优化了对变形链球菌的电击转化方案。     

杜仲对铜胁迫后杜氏盐藻恢复速率的影响

本研究通过检测不同浓度的杜仲药液对铜胁迫下的盐藻细胞数和叶绿素含量,来探讨杜仲对铜胁迫后杜氏盐藻恢复速率的影响。结果表明,适当浓度的杜仲药液会加快盐藻的恢复速率,其中5mg/ml的药液作用最为明显。     

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.