目录
关键词 II
Abstract III
引言
1 材料与方法 1
1.1 生物信息学分析 1
1.1.1 数据来源 1
1.1.2 ATACseq数据分析 1
1.1.3 RNAseq数据分析 2
1.1.4 ATACseq和RNAseq关联分析 2
1.2 分子实验 3
1.2.1 实验材料 3
1.2.2 植物总DNA提取 4
1.2.3 烟草验证系统载体构建 4
1.2.4 烟草瞬时表达验证目的序列片段功能 7
1.2.5 玉米验证系统载体构建 8
1.2.6 玉米原生质体提取、质粒对原生质体的转化 9
2 结果与分析 11
2.1 增强子的预测结果 11
2.1.1 样本信息 11
2.1.2 ATACseq数据质量控制 12
2.1.3 RNAseq数据质量控制 12
2.1.4 ATACseq数据比对分析 13
2.1.5 预测增强子序列 16
2.2 烟草中预测增强子的功能验证 17
2.2.1 预测增强子目的片段的获得 17
2.2.2 片段连接到表达载体 19
2.2.3 烟草侵染转化实验结果分析 21
2.3 玉米中预测增强子的功能验证 21
2.3.1 预测增强子目的片段的获得 21
2.3.2 片段连接到表达载体 22
3 讨论 23
3.1 基于染色质可及性的增强子预测的效率 24
3.2 不同时期染色质可及性变化探究增强子的活性 24
致谢 24
参考文献 24
基于ATACseq和RNAseq的增强子预测与验证
摘 要
玉米是一种拥有复杂基因组的重要粮食作物，对玉米染色质结构的认识将有助于我们对作物进行遗传改良。增强子作为一种重要的顺式调控因子，参与基因表达调控，影响许多重要的农艺性状。然而，目 *51今日免费论文网|www.51jrft.com +Q: ^351916072^ 
前玉米中仅鉴定出少量的增强子，因此非常有必要进行全基因组水平的增强子预测和鉴定。随着测序技术的进一步发展，染色质开放测序技术逐渐运用到植物中，为进一步解析了玉米的染色质结构提供了可能。本研究利用生物信息学手段，通过对玉米ATACseq和RNAseq数据的综合分析，将两种数据peaks均富集的区域定义为预测增强子，我们共预测出32个增强子。同时我们挑选出2个预测序列与GFP报告基因融合，并在烟草和玉米中瞬时表达，验证其具有增强子功能。本研究基于ATACseq和RNAseq数据对增强子的预测有助于我们对玉米染色质结构的全面认识，并为分子设计育种提供一定的理论基础。
ENHANCER PREDICTION AND VALIDATION BASED ON ATACSEQ AND RNASEQ
ABSTRACT
Maize is an important crop with a complex genome. The understanding of the chromatin structures in maize provides opportunities for crop improvement. As an important cisregulatory factor, enhancer regulates gene expression, and affects the agricultural traits of plants. However, only a few identified enhancers are available in maize at present, so it is very necessary to predict and identify enhancers at the whole genome level. With the further development of sequencing technology, assay for transposaseaccessible chromatin sequencing (ATACseq) is gradually applied to plants, make it possible to further analyze the structure of corn chromatin. In this study, bioinformatics was used to analyze ATACseq data and RNAseq data from corn. 32 sequences, screened from the overlaps of these two regions sets, are defined as predictive enhancers, then we selected 2 predictive sequences to be combined with the GFP reporting gene and expressed instantaneously in tobacco and corn in order to further determine the enhancer function of the sequences. In this study, the prediction of enhancers based on ATACseq and RNAseq data is helpful to our comprehensive understanding of the chromosome structure of corn and provides a theoretical basis for molecular design and breeding.

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