目录
摘要II
关键词II
AbstractIII
引言
引言（或绪论）1
1 文献综述1
1.1 细胞分裂素 1
1.2 细胞分裂素双元信号系统 2
1.3 细胞分裂素受体2
2 方案论证3
3 材料与方法4
3.1 番茄细胞分裂素受体基因的鉴定4
3.1.1 基因组及功能注释信息4
3.1.2 细胞分裂素受体基因的搜索4
3.1.3 保守结构域预测4
3.1.4 多序列比对和系统树构建4
3.1.5 染色体定位4
3.1.6 遗传参数计算4
3.2 番茄细胞分裂素受体基因表达分析4
3.3 CRISPR/Cas载体的构建4
3.4 番茄的转化5
3.5 突变体鉴定5
4 结果与分析5
4.1 番茄细胞分裂素受体基因的鉴定5
4.2 系统进化树和蛋白序列保守结构域分析5
4.3 番茄细胞分裂素受体基因的染色体分布6
4.4 番茄细胞分裂素受体基因的遗传参数计算6
4.5 番茄细胞分裂素受体基因的表达分析6
4.6 SlHK4基因CRISPR/Cas编辑载体的构建8
4.7 SlHK4突变植株的获得与鉴定9
5 讨论10
致谢11
参考文献12
番茄细胞分裂素受体基因的生物信息学分析和突变体构建
摘 要
本文旨在运用生物信息学方法对番茄细胞分裂素受体基因家族成员进行鉴定和分析，并通过CRISPR/Cas9基因编辑技术构建细胞分裂素受体的功能突变体，从而更好的研究和利用番茄细胞分裂素受体基因。通过对番茄基因组的分析，我们已鉴定得到与拟南芥细胞分裂素受体基因AHKs同源的3个基因，分别命名为SlHK2、SlHK3和SlHK4。系统进化树和保守结构域分析表明，SlHK2、SlHK3和SlHK4与拟南芥AHK2、AHK3、AHK4一一对应，且SlHKs基因在进化上偏 *51今日免费论文网|www.51jrft.com +Q: *351916072* 
保守。遗传参数计算结果显示，SlHKs经历了纯化选择，估计基因对发生分化的时间为48.58Mya。SlHKs的表达分析显示，相对于SlHK2和SlHK3，受体SlHK4在MicroTom的生殖器官花、果实、种子中有更高的表达，推测SlHK4基因更有可能与番茄花器官和果实发育调控有关。因此，运用CRISPR/Cas9基因编辑技术构建受体SlHK4的功能突变体slhk4。对所得的T0、T1代植株进行突变位点的测序鉴定，已获得T1代纯合突变体株系，为进一步研究其生长发育变化和SlHK4基因的功能奠定了基础。
BIOINFORMATICS ANALYSIS AND MUTANT CONSTRUCTION OF THE CYTOKININ RECEPTOR GENE FAMILY IN TOMATO
ABSTRACT
This article aims to use bioinformatics methods to identify and analyze members of the tomato cytokinin receptor gene family, and to construct functional mutants of the cytokinin receptor through CRISPR/Cas genome editing technology, so as to better study and utilize the cytokinin receptor genes in tomato. Through BLASTp searches in the tomato proteome, we have identified three genes homologous to the Arabidopsis cytokinin receptor genes AHKs, and named as SlHK2, SlHK3, and SlHK4, respectively. Phylogenetic and domain analyses show that SlHK2, SlHK3, and SlHK4 are orthologous to Arabidopsis AHK2, AHK3, and AHK4, respectively, and the SlHK genes are evolutionarily conservative. Ks and Ka/Ks values of the SlHK genes demonstrate that SlHKs has undergone purification selection, and their estimated divergence time is 48.58Mya. Expression analyses of SlHKs show that, compared with SlHK2 and SlHK3, the receptor gene SlHK4 has higher expression levels in the reproductive organs, including flowers, fruits and seeds, of MicroTom. Therefore, we speculated that the SlHK4 gene may play a more important role than the other two genes in the regulation of development of tomato flowers and fruits. We selected SlHK4 as the target gene, and constructed its functional mutants slhk4 using the CRISPR/Cas genome editing technology. We has identified homozygous mutant lines from the T1 generation plants, providing a basis for future studies on the changes in the growth and development of the mutant plants and on the roles of the SlHK4 gene in tomato.

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