辣椒果实细胞学、转录组分析及赤霉素受体CaGID1s和DELLA蛋白CaGAI功能、互作研究

发布时间：2018-06-19 04:13

本文选题：辣椒 + 果实　；参考：《中国农业大学》2015年博士论文

【摘要】：从一个辣椒RIL (recombinant inbred line,重组自交系)群体中选出果实大小和形状各异的49份材料,连同双亲及杂交一代共52份材料为试材,对子房和成熟果实进行细胞学研究发现,在辣椒果皮横切和纵切面上,果皮(子房壁)的结构很保守,在授粉之前已经确定,果皮细胞可以清晰的划分为：外果皮、中果皮和内果皮细胞。相关性分析表明,果实横径的生长模式比较保守；除细胞形状外,细胞数目也在很大程度上影响果实形状。从果实的宏观生长来看,果实横径与果皮厚度正相关,果实纵径与二者不相关,呈现出异速增长模式,形成了果实形状的多样性。果皮细胞的生长模式与果实宏观生长相似,也表现出异速增长模式。以羊角形辣椒自交系“0819”为试材,研究了果实生长过程中外部形态和细胞的动态变化。“0819”果实生长过程中,果实纵径、横径和果皮厚度具有相似的“S”形生长曲线,在纵向和横向切面上,中果皮细胞平均面积增长曲线也大致呈“S”形。细胞径向分裂主要发生在果实生长早期的最外层细胞,以增大果实横切面周长适应中果皮细胞的膨大；在10DAP (days after pollination,授粉后天数)之前,外果皮及中果皮细胞进行活跃的横向分裂,是果实纵径伸长的重要影响因素；平周分裂主要发生在外层亚表皮细胞层,大约在20DAP停止。细胞膨大主要发生在中果皮,并对果实横径、果皮厚度以及后期的果实纵径伸长起决定作用。采用ELISA方法,对0、5、10、50DAP果实中IAA、 GA、 ZR和ABA的含量进行测定,发现种子和胎座中激素含量的变化趋势大致相同,而果皮中的变化趋势与之有明显差异。在5DAP时辣椒种子中赤霉素含量升高,生长素含量降低,而在5DAA-NP时种子生长素含量仍然较高,赤霉素含量降低,说明赤霉素可能是促进辣椒果实坐果的主要激素,而且授粉受精后种子中赤霉素的合成并非与生长素的大量合成相关。对0、5、10、50DAP果实进行转录组测序,发现在5和10DAP专一表达的基因数目比0DAP和50DAP专一表达的基因数目少,说明5和10DAP果实主要进行膨大生长,生长状态相似,而0DAP和50DAP时,果实(子房)进行着更为复杂的反应；植物激素相关基因的表达也显示,5DAP和10DAP的果实基因的表达模式相似。同源克隆了3个赤霉素受体基因CaGIDlb.1、 CaGIDlb.2和CaGID1c,以及一个赤霉素信号转导途径中的负调控因子DELLA蛋白基因CaGAI。通过基因表达定量分析,发现在辣椒坐果过程中,CaGIDlb.2在果皮和胎座中发挥着重要作用；而CaGIDlb.／则在种子中可能有更重要的作用。在果实生长发育的过程中,CaGIDIb.l和CaGIDlb.2可能比CaGIDlc有更重要的作用。将CaGIDlb.1、 CaGIDlb.2和CaGID1c在拟南芥双突变体gidlagidlc中过表达,株高显著增加；将CaGAI在拟南芥双突变体rga-24/gai-t6中过表达,株高降低,表明CaGIDls和CaGA1影响植株茎的伸长,与拟南芥中AtGID1和AtGAI的作用相似。酵母双杂(Y2H)和荧光双分子互补(BiFC)试验表明,在GA3存在的条件下,CaGID1b1和CaGID1b2可以与CaGAI相互作用；而CaGID1c与CaGAI的相互作用不依赖于GA3。
[Abstract]:49 samples of different sizes and shapes of fruit were selected from a group of RIL (recombinant inbred line, recombinant inbred line). Together with 52 copies of a total of two parents and hybrids, cytological studies on the ovary and mature fruit found that the structure of the peel (ovary wall) was conservative in the cross and longitudinal section of Capsicum. Before the powder was determined, the peel cells could be clearly divided into the outer skin, the pericarp and the endocarp cells. The correlation analysis showed that the growth pattern of the transverse diameter of the fruit was more conservative; the cell number also influenced the shape of the fruit to a great extent except the shape of the cells. The longitudinal diameter of fruit is not related to the two, showing a pattern of different speed growth, which forms the diversity of fruit shape. The growth pattern of fruit skin cells is similar to the macro growth of fruit, and also shows a pattern of different speed growth.
The dynamic changes of external morphology and cells in the growth process of the capsicum pepper inbred line 0819 were studied. "0819" fruit growth process, the fruit longitudinal diameter, transverse diameter and pericarp thickness have similar "S" shape growth curve, and the average area growth curve of the pericarp cells in the longitudinal and transverse sections is also roughly presented. "S" shape. Cell radial division mainly occurs in the outermost cells of the early growth of the fruit, which increases the expansion of the pericarp cells in the transverse section of the fruit. Before the 10DAP (days after pollination, the number of days after pollination), the active transverse division of the outer skin and the medium pericarp cells is an important factor affecting the lengthwise elongation of the fruit. The division of the circumferential Division mainly occurs in the outer layer of the subepidermal cell layer and stops at about 20DAP. The cell expansion mainly occurs in the pericarp, which determines the transverse diameter of the fruit, the thickness of the pericarp and the lengthwise elongation of the later fruit.
ELISA method was used to determine the content of IAA, GA, ZR and ABA in the fruits of 0,5,10,50DAP. It was found that the change trend of hormone content in seed and placenta was approximately the same, but there was a significant difference in the pericarp. At 5DAP, the content of gibberellin and the content of auxin in the pepper seeds were lower, while the seed auxin was contained in 5DAA-NP. The amount of gibberellin may be the main hormone to promote fruit sitting of capsicum, and the synthesis of gibberellin in seed after pollination and fertilization is not related to the synthesis of auxin.
The transcriptional sequence of 0,5,10,50DAP fruit showed that the number of genes expressed in 5 and 10DAP was less than that of 0DAP and 50DAP, indicating that 5 and 10DAP fruits were mainly expanded, and the growth state was similar, while the fruit (Zi Fang) was more complex when 0DAP and 50DAP, and the expression of plant hormone related genes was expressed. It also showed that the expression patterns of 5DAP and 10DAP were similar.
Homologous cloning of 3 gibberellin receptor genes CaGIDlb.1, CaGIDlb.2 and CaGID1c, and a negative regulator of gibberellin signal transduction pathway DELLA protein gene CaGAI. through quantitative analysis of gene expression, it is found that CaGIDlb.2 plays an important role in the peel and placenta during the fruit setting of capsicum, and CaGIDlb. / in the seed. In the process of fruit growth, CaGIDIb.l and CaGIDlb.2 may have a more important role than CaGIDlc. CaGIDlb.1, CaGIDlb.2 and CaGID1c are overexpressed in the Arabidopsis double mutant gidlagidlc, and the height of the plant increases significantly; CaGAI is overexpressed in the Arabidopsis thaliana double mutant rga-24/gai-t6, and the height of the strain is reduced. The effect of CaGIDls and CaGA1 on the elongation of plant stems is similar to that of AtGID1 and AtGAI in Arabidopsis. Yeast double heterozygosity (Y2H) and fluorescent double molecular complementation (BiFC) test show that CaGID1b1 and CaGID1b2 can interact with CaGAI under the presence of GA3, while the interaction between CaGID1c and CaGAI is not dependent on the CaGAI.
【学位授予单位】：中国农业大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：S641.3

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