巨桉、毛果杨、雷蒙德氏棉LBD基因家族的研究

发布时间：2018-03-14 11:18

本文选题：巨桉　切入点：毛果杨　出处：《中国林业科学研究院》2017年博士论文　论文类型：学位论文

【摘要】：转录因子是功能基因组时代的研究热点。植物中的家族转录因子参与调控了许多重要的生物学过程,包括形态建成、信号转导、环境应激反应等。LBD(Lateral Organ Boundaries Domain)/ASL(ASYMMETRIC LEAVES2-LIKE)蛋白是一类植物所特有的转录因子,在调控植物生长发育、营养代谢以及逆境胁迫响应等方面发挥重要作用。巨桉和毛果杨是重要的速生木本植物,其木材被广泛用于木质纤维素生物燃料的生产、造纸与纸浆工业以及纤维素原料的产品。棉花是最重要的经济作物之一,它是纤维及油料的重要来源。自2006年、2012年、2014年毛果杨、雷蒙德氏棉、巨桉全基因组测序工作分别完成以来,使得在基因组水平上对LBD基因家族进行分析成为可能。因此,本文以巨桉、毛果杨、雷蒙德氏棉为研究对象,从中鉴定出LBD基因家族并成功筛选出一批重要的LBD基因,期望为今后促进木本植物木材的形成和提高其纤维产量提供基因资源和技术支撑。主要结果如下:利用生物信息学方法,从巨桉基因组中共鉴定出46个LBD基因,发现它们不均匀地分布在11条染色体上。基于进化关系分析,巨桉LBD基因可以分为ClassⅠ和ClassⅡ两大类,并进一步细分为Ⅰa、Ⅰb、Ⅰc、Ⅰd、Ⅰe、Ⅰf、Ⅱa和Ⅱb等8个亚类。通过qRT-PCR检测LBD基因家族成员在不同组织的表达模式,发现EgLBD29基因在木质部特异地高表达,EgLBD22和EgLBD37基因在韧皮部特异地高表达。共聚焦显微镜观察结果表明,EgLBD22、EgLBD29和EgLBD37蛋白定位于细胞核。此外,本研究还发现EgLBD22、EgLBD29和EgLBD37基因的表达响应生长素和赤霉素胁迫处理。通过挑选这三个巨桉LBD基因在杂交杨树种84K(Populus alba×Populus glandulosa)中异位表达研究其功能,结果证明这三个基因对84K次生生长过程产生了不同程度的影响:过表达EgLBD37基因的84K杨的次生木质部显著增厚,过表达EgLBD29基因的84K杨的韧皮纤维显著增多,过表达EgLBD22基因的84K杨则没有明显的表型变化。通过对EgLBD22-oe、EgLBD29-oe和EgLBD37-oe转基因84K和野生型84K进行转录组测序,发现转基因84K杨中一些与赤霉素、乙烯、生长素信号通路、细胞扩增相关的重要基因的表达量明显上调或下调。这些新发现表明EgLBD29和EgLBD37基因在巨桉次生生长过程中发挥着重要的作用,本研究得到了一种促进木材形成和提高纤维产量的新方法。从毛果杨基因组中鉴定出57个LBD基因并预测其分子量和等电点,结果表明:这些基因分布于毛果杨18条染色体以及Scaffold_65、Scaffold_86上。该家族成员基因结构简单,内含子数目不超过2个。进化关系分析显示毛果杨LBD基因可以分为ClassⅠ和ClassⅡ两大类,并进一步细分为Ⅰa、Ⅰb、Ⅰc、Ⅰd、Ⅰe、Ⅱa和Ⅱb等7个亚类。分析基因表达模式发现,该家族成员的表达具有一定时空特异性,并响应氮素胁迫处理。这些工作为今后深入研究毛果杨LBD基因的功能奠定了基础。从雷蒙德氏棉基因组中鉴定出66个LBD基因,发现它们不均匀的分布在13条染色体上。这些基因可以分为ClassⅠ和ClassⅡ两大类,细分为Ⅰa、Ⅰb、Ⅰc、Ⅰd、Ⅱa和Ⅱb等6个亚类。该家族基因结构简单,外显子数目不超过3个。从雷蒙德氏棉LBD家族成员中共找出15段保守基序,并对LOB(Lateral Organ Boundaries)结构域的保守性进行了分析。基因表达模式分析揭示出雷蒙德氏棉LBD基因家族的表达具有一定时空特异性,这些结果为今后雷蒙德氏棉LBD基因功能的验证奠定了基础。
[Abstract]:Transcription factor is a hot research topic in functional genomics era. The family of transcription factors in plants is involved in many important biological processes, including morphogenesis, signal transduction, environmental stress reaction.LBD (Lateral Organ Boundaries Domain) /ASL (ASYMMETRIC LEAVES2-LIKE) protein is a transcription factor specific to a class of plant growth and development, in the regulation of plant and play an important role in nutrient metabolism and stress response. Eucalyptus and poplar is an important fast-growing woody plants, the wood is widely used in the production of lignocellulosic biofuels, products of paper and pulp industry and cellulose raw materials. Cotton is one of the most important economic crops, it is an important source of fiber and oil. Since 2006, 2012, 2014 Raymond S p.trichocarpa, cotton, Eucalyptus grandis genome sequencing work were completed to make at the genomic level The LBD gene family of analysis possible. Therefore, in this paper, Eucalyptus grandis, Populus trichocarpa, Raymond S cotton as the research object, from the identified LBD gene family and successfully screened a number of important LBD gene, is expected to provide gene resources and technical support for the future promotion of the fiber yield of wood and woody plants to improve. The main results are as follows: using bioinformatics methods, from E. grandis genome identified 46 LBD genes and found that they are not uniformly distributed in 11 chromosomes. Based on the analysis of evolutionary relationship of Eucalyptus grandis LBD gene can be divided into Class I and Class II two categories, and further subdivided into 1 a. I B, I C, d I, e I, f I, II a II and B 8 subtypes. By qRT-PCR detection of LBD gene family expression pattern in different tissues, EgLBD29 gene was found in xylem specific high expression of EgLBD22 and EgLBD37 genes in the phloem. High specific expression. Confocal microscopy showed that EgLBD22, EgLBD29 and EgLBD37 protein located in the nucleus. In addition, the study also found that EgLBD22, in response to auxin and gibberellin stress expression of EgLBD29 and EgLBD37 gene. By selecting the Three Eucalyptus grandis LBD gene in hybrid poplar 84K (Populus Alba * Populus glandulosa) ectopic expression of its function, the results demonstrate that the three genes had different effect on the secondary growth process of 84K: the expression of EgLBD37 gene in poplar 84K secondary xylem thickening and the expression of EgLBD29 gene in 84K poplar phloem fiber was increased. The expression of EgLBD22 gene in poplar 84K had no obvious phenotypic changes. The EgLBD22-oe, EgLBD29-oe and EgLBD37-oe transgenic 84K and wild type 84K transcriptome sequencing, found in some transgenic poplar 84K and ethylene, gibberellin, auxin Signal pathway, cell expansion expression important genes related to the up-regulated or down regulated. These new findings show that play an important role in the growth of EgLBD29 and EgLBD37 genes in Eucalyptus grandis secondary process, this study got a promotion of wood formation and improve the new method of fiber production. From the identified 57 LBD genes in hair if the poplar genome and predict its molecular weight and isoelectric point. The results show that the distribution of these genes in Populus trichocarpa 18 chromosomes and Scaffold_65 Scaffold_86. The gene has the advantages of simple structure, intron number is less than 2. The evolutionary relationship analysis showed that LBD gene of Populus trichocarpa can be divided into Class and 1 Class II two categories, and further subdivided into 1 A, 1 B, 1 C, 1 D, 1 e, 2 A and B II 7 subtypes. Analysis of gene expression patterns showed that expression of members of the family has a certain temporal specificity and response to nitrogen stress . these work laid the foundation for further research on LBD gene of Populus trichocarpa function. From the identified 66 LBD gene Raymond cotton genome, see that they are not evenly distributed in 13 chromosomes. These genes can be divided into Class I and Class II two categories, subdivided into 1 A, 1 B, I C, I D, a II and B II 6 subtypes. The gene structure of the family, the exon number no more than 3. Find out 15 conserved motifs from the members of the Raymond S family and the cotton LBD, LOB (Lateral Organ Boundaries) domain of conservation are analyzed. Gene expression pattern analysis revealed the expression of Raymond S cotton LBD gene family has a certain temporal specificity, these results lay the foundation for future validation of Raymond S cotton LBD gene function.

【学位授予单位】：中国林业科学研究院
【学位级别】：博士
【学位授予年份】：2017
【分类号】：S792.11;S792.39;S562

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