Cloning and Expression of Genes from Venom Gland of Bungarus multicinctus

本研究利用前期研究中构建的pGEX-BgTX（P22-A31）质粒在大肠杆菌BL21（DE3）中表达并纯化后得到了较纯的重组α-银环蛇毒素同工毒素，得率约为1.225mg/L。用重组蛋白与天然α-银环蛇毒素的抗原性一致。重组α-银环蛇毒素同工毒素的半致死剂量LD50为1.598mg/kg，约为天然α-银环蛇毒素的1/5且有一定的镇痛药效，1/4LD50剂量的镇痛百分率为55.2%，1/8LD50剂量的镇痛百分率为20.5%，在外周镇痛作用中呈一定的量效关系。以上结果表明，本研究重组表达的α-银环蛇毒素同工毒素（P22-A31）具有与天然α-银环蛇毒素相似的生物学活性。α-银环蛇毒素cDNA序列...α-bungarotoxin plays very important role in neuroscience research, clinical application and the pharmaceutical industry. In order to acquire quantites of α-bungarotoxin, we expressed GST-α-bungarotoxin fusion protein using constructed plasmid pGEX-BgTX (P22-A31) in E. coli BL21 (DE3) cell and finally obtained recombinant α-bungarotoxin with yields of about 1.225 mg/L. The results of both ELISA and...学位：理学硕士院系专业：生命科学学院生物学系_生物化学与分子生物学学号：20032610

两个蛇毒基因克隆及cDNA序列多态性再分析

α-银环蛇毒素(α-bungarotoxin)是一种突触后神经毒素,广泛存在于眼镜蛇科蛇类的毒腺中,对于该基因cDNA多态性是否真实一直存有争议。本研究从银环蛇基因组DNA中克隆到α-银环蛇毒素基因序列,并对其中5个克隆进行测序和序列比对分析。作为参照,从同一次反转录得到的cDNA混合物中,克隆了蛇毒神经生长因子cDNA,并对其进行测序、比对和突变情况分析。综合各研究组报道的α-银环蛇毒素cDNA序列、α-银环蛇毒素基因序列和神经生长因子cDNA序列的突变情况,发现α-银环蛇毒素cDNA的多态性在基因组模板上不存在对应的变化,因此推测这种多态性不是从不同的转录本而来,同时考虑到不同研究小组报道的序列突变位点并没有出现相同的情况,因此其多样性也不是RNA编辑的结果。可见这种cDNA序列上的多样性很可能是由反转录过程以及基因克隆过程中人为引入的错误造成的

The Application of Bioinformatics in the Research of Alternative Splicing

基因选择性剪接现象是真核生物基本而又重要的调控机制。由于基因的选择性剪接在形成生物复杂性和多样性上具有极其重要的作用,同时选择性剪接与许多人类疾病也密切相关。因此,研究基因选择性剪接是一项十分重要的工作。生物信息学作为一门新兴的学科在研究基因选择性剪接上起关键的作用,尤其在研究基因表达调控机制、选择性剪接基因预测以及选择性剪接基因进化上。文章综述了这方面的最新研究进展,为更深入了解真核生物基因的表达调控机理提供依据。Alternative splicing,a fundamental and important regulatory mechanism in eukaryotes,allows one pre-(mRNA) to be processed into many different mature forms within a cell,each of which can have distinct functions.As alternative splicing is associated with human diseases,the study of alternative splicing becomes quite important.Bioinformatics is a new subject for the study of alternative splicing,especially for its regulatory mechanism,prediction and origin.Of course,bioinformatics must be combined with experimental research so as to clarify these aspects of alternative splicing.This paper reviewed the recent research progress in this field in the hope to gain a deeper understanding of eukaryotic gene expression regulation.国家自然科学基金项目(编号:60305001,30470938)资助~

Expression and functional analysis of recombinant α-bungarotoxin (P22-A31)

α-银环蛇毒素在神经科学研究以及在临床和制药上有重要作用,为获得大量的α-银环蛇毒素,我们用已构建的pGEX-BgTX(P22-A31)质粒在大肠杆菌BL21(DE3)中表达,并用谷胱甘肽Sepharose FF纯化GST-α-银环蛇毒素融合蛋白,再用凝血酶切掉融合标签谷胱苷肽转移酶(Glutathione S-transferase,GST),得到了较纯的重组α-银环蛇毒素同工毒素,得率约为1.225mg/L。用重组α-银环蛇毒素制备多克隆抗体,经ELISA和Western杂交鉴定后可知重组α-银环蛇毒素与天然α-银环蛇毒素的抗原性一致。小鼠毒性试验表明,重组α-银环蛇毒素同工毒素的半致死剂量LD50为1.598mg/kg,约为天然α-银环蛇毒素的1/5。小鼠化学法镇痛试验显示,重组α-银环蛇毒素有一定的镇痛药效,1/4LD50剂量的镇痛百分率为55.2%,1/8LD50剂量的镇痛百分率为20.5%,在外周镇痛作用中呈一定的量效关系。α-bungarotoxin plays very important role in neuroscience research, clinical application and the pharmaceutical industry. In order to acquire quantitites of α-bungarotoxin, we expressed GST-α-bungarotoxin fusion protein using constructed plasmid pGEX-BgTX (P22-A31) in E.coli BL21 (DE3)cell, purified GST-α-bungarotoxin using glutathione-Sepharose FF, cleavaged Glutathione S-transferase(GST) tag by thrombin in the column, and finally, we obtained recombinant α-bungarotoxin with yields of about 1.225 mg/L. In order to compare the immunogenicity of recombinant protein and crude α-bungarotoxin, we prepared multi-antibody using recombinant α-bungarotoxin. The results of both ELISA and Western blot showed that recombinant α-bungarotoxin has the same antigenicity as natural α-bungarotoxin. In vivo toxicity tests showed that the LD_ 50 of recombinant α-bungarotoxin was 1.598 mg/kg, about 1/5 that of natural α-bungarotoxin. Analgesis percentages with doses of 1/4 LD_ 50 and 1/8 LD_ 50 were 55.2% and 20.5% respectively, indicating that recombinant α-bungarotoxin possesses analgesic efficiency.国家自然科学基金(No.30470938);; 教育部留学回国人员启动基金资助~

The Phylogeny Tree Reconstruction Based on the Usage Frequencyof Codons and Corresponding Complementary Codons

为了更深入地了解密码子与互补密码子使用相关的内涵,70个单细胞和多细胞生物被更进一步的分析。结果得到70种细胞生物具有较高的相关系数平均值(0.5101,标准差是0.1389),表明密码子与其互补密码子使用间确实存在正的使用相关联系,进一步地支持了以前的结果。此外,通过对70个单细胞和多细胞生物在基于密码子与互补密码子使用频率差异,以及配对频率的高低信息的分析基础上,绘制进化树,得到的进化树与传统的进化树非常接近。The hypothesis that a codon bias usage is identical to its complementary codon usage preference has been investigated by using the relationship analysis of codons vs their complementary ones among 70 organisms. Significantly positive usage correlations between codons and their complementary ones were found and its implication in biology was also analyzed. The codoncomplementary codon tree was further built, which fairly exhibited the evolutionary relationship of these organisms. The results not only demonstrated the validity of our hypothesis,but also manifested the usefulness of correlation analysis in studying on codon usage pattern and molecular evolutionary mechanisms of organism.ThisworkwassupportedbyagrantfromtheNationalNaturalScienceFoundationofChina(No.60305001)andagrantfromtheScienceandTechnologyMinistryofChina(PreliminaryStudyonFunctionalGenomeSystematics)(No.2001CCA01400)

Correlation Analyses of Codons and Corresponding Complementary Codons Usage

遗传密码是基因与蛋白质进行信息对接的桥梁,研究不同进化阶层生物的密码子与互补密码子的使用规律是揭示物种进化的一种重要途径。为了阐明不同生物和细胞器基因密码子与互补密码子使用之间的内在联系及其在进化中的意义,作者选取了44种细胞生物、16种病毒、40种线粒体和20种叶绿体的基因密码子与互补密码子数据,并进行关联分析。结果显示:(1)无论是古细菌,真细菌,病毒,真核生物还是两类细胞器,它们的密码子与互补密码子的使用频率大多表现为显著或极显著正相关;(2)不同生物和细胞器基因密码子与互补密码子使用频率之间相关程度也存在明显的异质性。细胞生物和两类细胞器基因密码子与互补密码子使用的高度一致性反映了它们可能具有共同的起源,显著的异质性则反映了它们分别经历了不同的进化选择压力。密码子与互补密码子的使用相关程度不仅能被用作为测量物种进化过程中所经历选择压力的指标,而且也能被用作为碱基摆动和修饰的指示器。此外,文中还对生物进化早期存在DNA双链同时具有编码基因功能的可能性进行了讨论。To reveal the relationship between codons and corresponding complementary codons usage, the data of codon and corresponding complementary codon usage from 44 cellular organisms, 16 viruses, 40 mitochondria and 20 chloroplasts were analyzed. Significant correlations between codons and corresponding complementary codons usage were found in the majority of the studied subjects except viruses, albeit there was also significant heterogeneity among them. The degree of correlation between codons and corresponding complementary codons usage may be used as an index not only for measuring the degree of selection force during organism evolution, but also for exploring the degree of anticodons' modification and wobbling-pairing with corresponding codons within organisms. In addition, it is also discussed that the two strands of DNA may have simultaneous coding function in the early stage of organism evolution.国家自然科学基金项目((No 60305001)资助

苹果蠹蛾对新疆林果业发展影响的评估及风险管理[J]

于2012年7月对新疆地区林业有害生物苹果蠹蛾进行系统调查研究，以便为新疆林业有害生物苹果蠹蛾的危险性进一步研究提供重要参考依据。在实际调查中发现，苹果蠹蛾是一种危害苹果、梨、杏等多种果实的入侵物种且危害较重，采用风险评估方法研究苹果蠹蛾对新疆林果业发展的影响。结果显示，苹果蠹蛾在新疆符合检疫性有害生物的地理分布管理标准，新疆具有苹果蠹蛾生存与蔓延扩散的环境条件，是新疆林果业的重要有害生物，该害虫具有在新疆境内蔓延扩散并暴发成灾的潜在风险，对新疆林果业发展构成了严重威胁。因此，应控制该虫扩散蔓延传入危害发生，并针对苹果蠹蛾提出监测预防和检疫管理措施，为新疆林果业经济发展提供参考

喀什旅游文化产业发展模式与保障机制研究[C]

文化与旅游的相互交融性,决定了文化产业与旅游产业结合是当今发展的重要趋势,受到各国各地区旅游业界和学界的普遍关注.本文探讨了旅游文化产业的构成要素、影响因素及特征,结合旅游评价原则和方法,构建了旅游文化产业评价指标体系.新疆喀什地区发展旅游文化具有得天独厚的优势,但目前,喀什地区旅游文化产业发展潜力尚未充分体现

喀纳斯图瓦村落社区参与旅游发展调查/Community participation in tourism development of Tuva Village at Kanas District[J]

采用问卷调查和深度访谈相结合的方法,调查喀纳斯景区及周边三个图瓦村落旅游开发十余年来的社区参与情况,分析得出:(1)图瓦居民社区参与旅游的类型属于“被动参与”型,以个体经营为主；(2)图瓦居民普遍表达了对旅游发展的积极态度,对旅游培训和就业表现出很强的参与意愿,但在旅游投资、规划、决策和景区管理方面参与意识淡薄；(3)图瓦村产值结构实现了由牧业收入主导向旅游收入主导的转变,劳动力也逐步由畜牧业向旅游业转移；(4)图瓦文化内涵挖掘与保护不足,低水平开发泛滥.基于上述研究结果,提出了提高图瓦居民社区参与度的对策建议

新疆喀什民族文化旅游开发模式评价/Evaluation of development models for ethnic cultural tourism in Kashgar City, Xinjiang[J]

通过文献分析法和专家咨询,在分析和总结国内外已有文化旅游开发模式的基础上,运用层次分析法构建了喀什民族文化旅游开发模式评价指标体系,并确定了各评价指标的权重；对喀什市、疏附县、疏勒县构建的“喀什金三角”14个文化旅游景区(点)进行评价,选择出各景区(点)最优开发模式；根据评价结果探讨了6种开发模式的内涵,提出了各景区(点)文化旅游开发的对策和建议.由于采用定性分析与定量分析相结合的方法,提高了研究结果的准确性,为喀什民族文化旅游开发模式选择提供借鉴