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1.
The influence of factors on the substrate-specificity of Pst Ⅰ restriction endonuclease has been studied with the method of electrophoresis. The results show that, the specificity of Pst Ⅰ almost can not be influenced by the single alteration of the concentration of Tris*HCl, Mg2+ or Na+ in the reaction system, but it can be altered by the reduction of any two of them. The specificity can not be altered by the single alteration of pH or the replacement of Mg2+ with Mn2+. The addition of glycerol or dimethylsulphoxide (DMSO) to the reaction system results in the relaxation of the substrate-specificity of Pst Ⅰ , but dimethylmethylformide, glycol and ethyl alcohol can not bring about the alteration of Pst Ⅰ specificity. Through the method of cloning and sequencing, the nucleotides of No.1 and 6 in the recognition sequence of Pst Ⅰ have changed (1C→A or 6G→T). Used with the enzyme analysis of an artificially synthetic DNA segment containing a special sequence, the nucleotides of No.1 and 6 have both changed (1C→A and 6G→T). The recognition sequence of Pst Ⅰ is speculated to be changed from CTGCA↓G to TGCA↓.  相似文献   

2.
The irreversible modifying effects onPst I of several inhibitors have been studied with the irreversible inhibition kinetic theory of single substrate reaction provided by Tsou, C. L. Pyridoxal phosphate (PLP), p-chloromercuribenzoic acid (PCMB), diisopropyl fluorophosphate (DEP), 2,3-diacetyl (DAC) and N-ethyl-5-phenylisoxazoliun-3′-sulfonate (woodward's reagent K, WRK) modify the lysine, cysine, serteine, arginine and carboxyl groups of the protein molecule respectively. These five inhibitors have been found to inhibit both the prime activity and star activity ofPst I. Used with the irreversible inhibition theory, the apparent inhibition rate constant,A and the microcosmic inhibition rate constants,k +0 andk′ +0 of every inhibitor were calculated. We also found that their inhibition effects belong to the noncompetitive irreversible inhibition. Results show that among the groups to be modified, some have nothing to do with the combination with the substrate, and some may have, but any of them isn't the only factor involved in the specific binding. Despite all this, they may take part in the catalysis of enzyme or have important effects on maintaining the active structure of enzyme molecules. Furthermore, serine and arginine residues are related to the alteration ofPst I conformation and then influence the ability ofPst I recognizing and incising DNA specifically. Foundation item: Supported by the Research Fund for the Doctoral Program of Higher Education (1999048601) Biography: Zou Guo-lin (1947-), male, Professor, research direction: enzymology.  相似文献   

3.
In 1991, we found that methylation outside the Pvu II recognition sequence could partially inhibit its cleavage activity. To clarify the molecular mechanism, three plasmids with different methylation states were constructed. Then, together with the original one, four plasmids were digested with different amounts of Pvu II. Results show that methylation on both sites results in 90 % inhibition; moving the methylated site one base further away decreases the inhibitory effect to about 30 %; with the adjacent dam methylation site eliminated, the inhibitory effect disappears. The data suggest that the inhibition of cleavage activity caused by outside methylation is not “all or none”, and the degree of inhibition is dependent on the position and the number of methylated bases.  相似文献   

4.
刘隽  高成卓  邹国林 《河南科学》2009,27(9):1077-1081
分别以N-溴代琥珀酰亚胺(NBS),2,3-丁二酮和N-乙基-5-苯基异口唑-3’-磺酸盐作修饰剂,研究色氨酸,精氨酸残基和羧基对RtⅠ活性的影响.结果表明,NBS的修饰虽可导致Pst Ⅰ活性丧失,但紫外吸收光谱和SDS-PAGE研究结果表明色氨酸残基并非Pst Ⅰ的活性基团,而精氨酸残基和羧基的修饰可导致PstⅠ和Rst Ⅰ活性丧失,并按Keech和Farrant动力学方法分析得出无论在原活性还是星号活性条件下,有一个精氨酸残基和两个羧基是Pst Ⅰ的活性基团.  相似文献   

5.
从脂肪嗜热芽孢杆菌FH58和S183中分别分离到Ⅱ型的限制性内切酶BstFⅠ和BstSⅠ。经鉴定,BstFⅠ的识别顺序和酶切位点为A/AGCTT,与Hind Ⅲ一致;BstSⅠ的识别顺序和酶切位点为C/PyCGPuG,与AvaⅠ一致。经肝素琼脂糖凝胶提纯,每克湿菌体可分离到10000单位的BstFⅠ或24000单位的BstSⅠ。BstFⅠ在45℃中保温6h,比活力不变,在50℃中保温1h比活力下降50%;而BstSⅠ在55℃中保温6h,比活力不变,在70℃中保温1h,比活力仅下降10%。  相似文献   

6.
7.
本文对我们构建的含有青霉素酰化酶(Acylace)基因的重组质粒pPAHD1进行了限制性内切酶图谱的测绘。使用了限制酶14种,证明BglⅡ,SalⅠ,SmaⅠ和BamHⅠ在重组质粒上各有切点一个;EcoRⅠ有切点两个:HindlⅡ,AvaⅠ各有切点三个;PvuⅠ有切点四个;EcoRⅤ和XmnⅠ各有切点五个。  相似文献   

8.
A theory of reverse triple I method of restriction for implication operator R0 is proposed.And the general computation formulas of infimum for fuzzy modus ponens and supremum for fuzzy modus tollens of a-reverse triple I method of restriction are obtained respectively.  相似文献   

9.
纯化至电脉均一纯的限制性核酸内切酶Bsp63I经SephadexG-200测得相对分子质量约为113800,SDS-聚丙烯酰凝胶电泳测得其亚基相对分子质量约为56800,用DNS法测得该酶N-末端氨基酸为丙氨酸,表明该酶由2个相同的亚基组成,还对Bsp63I和它的同工酶PstI和Bsp78I之性质进行了比较。  相似文献   

10.
Based on the theory of sustentation degree of Triple I method together with the formulas of a-Triple I modus ponens (MP) and a-Triple I modus tollens( MT), the theory of restriction degree of Triple I method is proposed. Its properties are analyzed, and the general formulas of supremum of a-Triple IMP and infimum of a-Triple I MT are obtained.  相似文献   

11.
限制的语言表达形式有三种:概念加限制词;不加限制词,属概念直接过渡到种概念;通过语境进行限制。概念的逻辑限制与语词的修饰从外延上看,二者是交叉关系,其中非限制性的修饰和定语性限制区别在于:非限制性的语词修饰,其修饰词不能缩小被修饰的语词所表达的概念的外延,定语性限制中的限制词即定语一定能缩小概念的外延;定语性限制的定语有区别性,非限制性修饰中的修饰词一般没有区别性,或者说修饰词的区别性不同于限制词的区别功能。  相似文献   

12.
给出图G和一个正整数r,令f‘r(G)为图G边染色的最大色数,使得每个顶点最多关联r种颜色,并且每个顶点关联的颜色中任两种颜色所染的边数相差最多为1.对所有的正整数n和r,给出了f‘r(Kn,n)的下界和上界;在r|n和r=2,3,n-1的情形,得到f‘r(Kn,n)的值。  相似文献   

13.
用 6 种限制性内切酶分析了中国白兔的线粒体 D N A(m t D N A)。测得其相对分子质量约为16.8, Eco RⅤ, Bam HⅠ, PstⅠ, Eco RⅠ, HindⅢ在中国白兔 m t D N A 上分别有 1,2,2,2,6 个切点, SalⅠ 在其上没有切点。根据单酶降解和双酶降解片段的相对分子质量,构建了中国白兔m t D N A的限制性内切酶图谱。  相似文献   

14.
油桐尺蠖核型多角体病毒(BsNPV)、木(木尞)尺蠖核型多角体病毒(CpNPV)和用CpNPV感染油桐尺蠖幼虫面得到的病毒CpNPV-Bs,三者经HaeⅢ酶解后的电泳图谱均一致,经HindⅢ酶解后,均获得11条片段。各病毒DNA的总分子量约为57.5×10~6D。本文初步认为BsNPV和CpNPV为同一种病毒。  相似文献   

15.
从喜温蓝藻SynechococuselongatusT3分离出一种限制性内切核酸酶,命名为SelI.此酶是限制性内切核酸酶Sau96I的同切酶,它们的识别序列GGNCC.但是SelI的反应条件与Sau96I不同.SelI是一种热稳定的限制性内切核酸酶,最适反应温度的范围是50~57℃  相似文献   

16.
:强化对权力的监督制约是巩固党的执政地位的要求 ,是保证权力正常运行的要求 ,也是加强党风廉政建设的根本所在。目前 ,对权力监督不力的问题较为突出。因此 ,必须匡正权力运行的轨道 ,对权力实行事前、事中、事后的全过程的监督制约  相似文献   

17.
从山东鄄城味精厂周围环境和发酵裂解液中分离得到三株噬菌体(Tp—1、Tp—2、Tp—3).电镜观察表明三株噬菌体在形态上是相似的,头部呈多面体,尾部细长,无鞘,属于长尾噬菌体类.DNA 的 EcoR Ⅰ和 Pst Ⅰ酶切分析结果表明在噬菌体 Tp—1和 Tp—2之间有一定差异.同时对噬菌体的寄主范围,感染特性,热稳定性和 pH 稳定性作了分析.  相似文献   

18.
经0.1%鸭血烯酸钠处理或经37℃溶菌酶作用的Bacillus subtilis BD366菌体,按碱法提取pUB110 DNA,可得到SC(超螺旋),OC(开环)和CT(Com-pact type,紧密型)结构的分子。SO和OC分子均可被EcoRⅠ,BamHⅠ和BglⅡ切成线状分子,但CT分子却不能。对pUB110DNA具有多切点的限制性内切酶Sau 3AⅠ对CT分子也不起作用。经86℃保温或沸水浴处理,CT分子能转变成SC和OC型,并可被Sau 3AⅠ酶切降解,推断CT结构可能是质粒的原始构型。  相似文献   

19.
将经San 3A部分酶切后的小麦总DNA片段克隆到λCharon40载体上构建了小麦基因文库.以两个人工合成的同源于小麦高分子量(HMW)谷蛋白基因的片段对此基因库进行筛选,获得了两个阳性克隆,推测其中一个包含了HMW谷蛋白全部的基因顺序.两个克隆的部分限制性内切酶位点已被确定.在Southern杂交中,用位于基因编码区内的寡核苷酸片段作探针,将阳性部分定位在一定的酶切片段上.  相似文献   

20.
鲍丽娟 《长春大学学报》2002,12(1):59-61,80
汉语中某些词语除了具有表面显现的反映义以外,还有隐含在使用上的某些特殊要求和含义。词语的隐义在其使用中表现为感情褒贬的不同,使用范围及语义轻重的差异,搭配上对词语的限定等。  相似文献   

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