RNA 序列分析 (1) 张高川生物信息学系 (Department of Bioinformatics) 基础医学与生物科学学院 (School of Biology and Basic Medical Science) 苏州大学医学部 (Medical College of Soochow Un

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1 RNA 序列分析 (1) 张高川生物信息学系 (Department of Bioinformatics) 基础医学与生物科学学院 (School of Biology and Basic Medical Science) 苏州大学医学部 (Medical College of Soochow University) Tel:

2 RNA 结构预测工具?

gene prediction software Family specific gene prediction software RNA homology search software

4 Single sequence secondary structure prediction Single sequence tertiary structure prediction Comparative methods Inter molecular interactions: RNA-RNA, MicroRNA:any RNA, MicroRNA:UTR ncrna gene prediction software Family specific gene prediction software RNA homology search software Benchmarks Alignment viewers/editors Inverse Folding/RNA design Secondary structure viewers/editors

36 怎么使用这些 RNA 结构预测工具?

38 Minimum free energy prediction Thermodynamic ensemble prediction

39 Minimum free energy prediction Thermodynamic ensemble prediction 5 6 Guide Tree based on multiple sequence alignment by ClustalW2 (

40 怎么计算 RNA 序列自身内部的碱基配对折叠?

41 密码子偏好性和优化问题?

42 Codon usage bias Codon usage bias refers to differences in the frequency of occurrence of synonymous codons in coding DNA.

43 Codon usage bias Effect on RNA secondary structure Because secondary structure of the 5 end of mrna influences translational efficiency, synonymous changes at this region on the mrna can result in profound effects on gene expression. Codon usage in noncoding DNA non-coding regions can therefore play a major role in RNA secondary structure and downstream protein expression, which can undergo further selective pressures. In particular, strong secondary structure at the ribosomebinding site or initiation codon can inhibit translation, and mrna folding at the 5 end generates a large amount of variation in protein levels.

44 Codon usage bias Effect on transcription/gene expression (I) Heterologous gene expression is used in many biotechnological applications, including protein production and metabolic engineering. Because trna pools vary between different organisms, the rate of transcription and translation of a particular coding sequence can be less efficient when placed in a non-native context. For an overexpressed transgene, the corresponding mrna makes a large percent of total cellular RNA, and the presence of rare codons along the transcript can lead to inefficient use and depletion of ribosomes and ultimately reduce levels of heterologous protein production. However, using codons that are optimized for trna pools in a particular host to overexpress a heterologous gene may also cause amino acid starvation and alter the equilibrium of trna pools.

45 Codon usage bias Effect on transcription/gene expression (II) This method of adjusting codons to match host trna abundances, called codon optimization, has traditionally been used for expression of a heterologous gene. However, new strategies for optimization of heterologous expression consider global nucleotide content such as local mrna folding, codon pair bias, a codon ramp or codon correlations. Specialized codon bias is further seen in some endogenous genes such as those involved in amino acid starvation. For example, amino acid biosynthetic enzymes preferentially use codons that are poorly adapted to normal trna abundances, but have codons that are adapted to trna pools under starvation conditions. Thus, codon usage can introduce an additional level of transcriptional regulation for appropriate gene expression under specific cellular conditions.

46 Codon usage bias Effect on speed of translation elongation Generally speaking for highly expressed genes, translation elongation rates are faster along transcripts with higher codon adaptation to trna pools, and slower along transcripts with rare codons. This correlation between codon translation rates and cognate trna concentrations provides additional modulation of translation elongation rates, which can provide several advantages to the organism. Specifically, codon usage can allow for global regulation of these rates, and rare codons may contribute to the accuracy of translation at the expense of speed.

47 Codon usage bias Effect on protein folding Protein folding in vivo is vectorial, such that the N-terminus of a protein exits the translating ribosome and becomes solvent-exposed before its more C-terminal regions. As a result, co-translational protein folding introduces several spatial and temporal constraints on the nascent polypeptide chain in its folding trajectory. Because mrna translation rates are coupled to protein folding, and codon adaption is linked to translation elongation, it has been hypothesized that manipulation at the sequence level may be an effective strategy to regulate or improve protein folding. Several studies have shown that pausing of translation as a result of local mrna structure occurs for certain proteins, which may be necessary for proper folding. Furthermore, synonymous mutations have been shown to have significant consequences in the folding process of the nascent protein and can even change substrate specificity of enzymes. These studies suggest that codon usage influences the speed at which polypeptides emerge vectorially from the ribosome, which may further impact protein folding pathways throughout the available structural space.

48 Codon Usage Database

49 Codon Usage Optimization

50 Codon Usage Optimization

55 RNA 种类和结构特征各类 RNA 数据库各类 RNA 分析工具 RNA 结构分析案例密码子偏好与优化学会灵活运用

56 课堂练习一 RNA 结构比较 (1) 同一个基因家族的不同成员 (2) 不同物种的同源基因 (3) 同一个 mrna 突变前后不少于三个

57 课堂练习二 mirna 靶标分析 (1) 通过 mirbase 挑选任意一个 mirna (2) 使用第三方的分析工具对该 mirna 的靶标进行分析

58 课堂练习三密码子优化改造 (1) 目标基因 : 人的任意一个编码蛋白的基因 (2) 目标物种 : 一是酵母 (yeast), 二是细菌 (E.coli)

59 拓展阅读和实践练习搜索并阅读有关 RNA 结构和功能的文献资料尝试练习 RNA 折叠相关分析工具的使用搜索并阅读密码子偏好性和优化方面的文献资料尝试练习密码子优化软件的使用

Microsoft Word - TIP006SCH Uni-edit Writing Tip - Presentperfecttenseandpasttenseinyourintroduction readytopublish

Microsoft Word - TIP006SCH Uni-edit Writing Tip - Presentperfecttenseandpasttenseinyourintroduction readytopublish 我难度 : 高级对们现不在知仍道有听影过响多少那次么 : 研英究过文论去写文时作的表技引示巧言事 : 部情引分发言该生使在中用过去, 而现在完成时仅表示事情发生在过去, 并的哪现种在时完态成呢时? 和难过道去不时相关? 是所有