【作者】 王冰；

【导师】 巩彪； 刘杰；

【作者基本信息】 山东农业大学 ， 农艺与种业（专业学位）， 2022， 硕士

【摘要】 番茄（Solanum lycopersicum L.）是世界范围内种植最广泛、消费最多的蔬菜之一。在生产过程中植物对磷肥的利用率低,限制了作物产量,造成了环境污染。通过采用具有磷高效基因的番茄植株作为砧木进行嫁接,可以提高植物在磷缺乏的环境中正常生长的能力,目前尚缺乏对磷高效基因的鉴定及其在农业生产中的应用。本研究采用“895自交系”、SAMS1过表达“SAMS1-OE”系为主要试验材料,在温室中采用霍格兰营养液进行培养,以正常磷（NP,1 m M H₂PO₄^-）霍格兰营养液作为对照,以低磷（LP）水培液中含有5μM H₂PO₄^-为处理组,待植株长到四叶一心后换LP水培培养。通过对番茄植株的根冠比、磷含量及利用指标、磷吸收、分配以及再利用等指标的测定,获得结果如下:SAMS1是一个PSR基因,通过酵母单杂交试验确认了PHR1与SAMS1启动子的相互作用,SAMS1是一个磷高效基因。在NP处理下,WT和SAMS1-OE植株在表型、幼苗生物量和根冠比差异不大;但是在LP胁迫12 d后,WT和SAMS1-OE植株均表现出生长抑制和根冠比增加的现象。但与WT相比,SAMS1-OE植株的地上部生物量较低,但根系生物量较高,导致植株的根冠比较高。通过使用多胺（PAs）和乙烯（Eth）抑制剂（MGBG和AOA）处理后发现,施用MGBG后在一定程度上抑制了SAMS1诱导的根生长,施用AOA后SAMS1抑制的地上部生长被恢复,诱导的根系生长被消除,这些结果表明过表达S AMS1通过PAs和Eth协同信号促进植物根系生长。在LP处理下,通过测定植物中Put、Spm、Spd的含量以及观察H₂O₂的代谢情况,发现SAMS1对PSR（磷饥饿反应）中PAs的代谢具有重要作用。LP处理下,SA MS1-OE会促进Put向Spm、Spd转化,保持了PSR过程中总PAs和（Spd+Spm）/Put比值的稳态。通过对植株施加外源抑制性药物,发现LP诱导的Put积累及其代谢的H₂O₂对根系的生长具有抑制作用。通过分析了PSR中Eth信号合成和转导突变体rin和nor植株的RSA（根系构型）,发现在NP处理下,植株的根系发育差异不大;但是在LP处理下,nor和rin突变体植株的侧根数量显著少于WT植株。这说明SAMS1诱导的Eth对侧根的生长具有促进作用,但是对根系的伸长影响不大。通过对植物中根以及茎叶中磷浓度、磷分配和磷吸收的情况分析,发现SAMS1-OE植株茎叶、根部磷含量及全株磷吸收均显著高于WT植株,过表达SAMS1降低了磷在茎叶中的分布,增加了磷在根部的分布;以地上部和地下部中的可溶性磷、细胞壁磷、果胶、果胶甲基酯酶作为评价磷素再利用的标准,发现LP胁迫导致两种形式的磷降低,但SAMS1-OE使植物中细胞壁磷的含量降低,可溶性磷的含量得到了提高。过表达SAMS1增加了两种组织的果胶浓度以及提高了果胶甲基酯酶的活性。为了进一步研究SAMS1在番茄全生育期对矿质营养吸收利用的情况,为此我们采用嫁接的方式,发现在LP处理下,用SAMS1-OE作为砧木植株的地上部干重和果实产量呈逐渐增加的趋势,根冠比高于WT植株,同时增施磷肥促进了磷从枝、根向果实的分配。以肥料利用率和产量贡献分析SAMS1对磷肥吸收利用的影响,使用转基因砧木提高了植株的吸磷量,PUE与施磷量呈负相关,在转基因砧木中磷肥的贡献比例显著降低,随着施磷量的增加,SAMS1对产量的贡献逐渐减小。以上结果表明,SAMS1可以促进Put的降解,促进侧根生长;Eth可以促进地上部细胞壁磷水解为可溶性磷,增加根部磷浓度,提高土壤中Apase活性,增加土壤中有机磷的释放,提高植物对磷元素的吸收;通过田间栽培发现,SAMS1作为砧木进行嫁接,可以提高磷肥利用率,提高番茄产量。为实际农业应用中提高磷肥利用效率提供了理论和技术支撑。更多还原

【Abstract】 Tomato（Solanum lycopersicum L.）is one of the most widely grown and consumed vegetables worldwide.In the production process,the utilization rate of phosphorus fertilizer by plants is low,which limits the crop yield and causes environmental pollution.Grafting tomato plants with phosphorus-efficient genes as rootstocks can improve the ability of plants to grow normally in phosphorus-deficient environments.At present,there is a lack of identification of phosphorus-efficient genes and their application in agricultural production.In this study,the"895 inbred line"and the SAMS1-overexpressing"SAMS1-OE"line were used as the main experimental materials,and then were cultured in Hoagland nutrient solution in the greenhouse,and normal phosphorus（NP,1 m M H₂PO₄^-）Hoagland was used for culture.The nutrient solution was used as a control,and the low-phosphorus（LP）hydroponic solution containing 5μM H₂PO₄^-was used as the treatment group,and the LP hydroponic culture was changed after the plants grew to four leaves and one heart.Through the determination of the root-shoot ratio,phosphorus content and utilization indicators,phosphorus absorption,distribution and reuse of tomato plants,the results are as follows:SAMS1 is a PSR gene,and the interaction of PHR1 with the promoter of SAMS1,a phosphorus efficient gene,was confirmed by yeast one-hybrid assay.Under NP treatment,WT and SAMS1-OE plants showed little difference in phenotype,seedling biomass and root-shoot ratio;but after LP stress for 12 d,both WT and SAMS1-OE plants showed growth inhibition and increased root-shoot ratio.Phenomenon.However,compared with WT,SAMS1-OE plants had lower shoot biomass but higher root biomass,resulting in a higher root-cap ratio of the plants.By treatment with polyamines（PAs）and ethylene（Eth）inhibitors（MGBG and AOA）,it was found that SAMS1-induced root growth was inhibited to some extent after MGBG application,and SAMS1-inhibited shoot growth was restored after AOA application,Induced root growth was abolished,and these results suggest that overexpression of SAMS1 promotes plant root growth through synergistic signaling of PAs and Eth.Under LP treatment,by measuring the contents of Put,Spm and Spd in plants and observing the metabolism of H₂O₂,it was found that SAMS1 plays an important role in the metabolism of PAs in PSR（phosphorus starvation response）.Under LP treatment,SAMS1-OE promoted the transformation of Put to Spm and Spd,maintaining the steady state of total PAs and（Spd+Spm）/Put ratio during PSR.By applying exogenous inhibitory drugs to plants,it was found that LP-induced Put accumulation and its metabolized H₂O₂ had inhibitory effects on root growth.By analyzing the RSA（root system configuration）of the rin and nor mutants in the synthesis and transduction of Eth signal in the PSR,it was found that under the NP treatment,the root development of the plants did not differ much;Plants had significantly fewer lateral roots than WT plants.This indicated that Eth was induced by SAMS1 to promote lateral root growth,but had little effect on root elongation.By analyzing the phosphorus concentration,phosphorus distribution and phosphorus absorption in roots,stems and leaves of plants,it was found that the phosphorus content in stems,leaves,roots and whole plant phosphorus absorption of SAMS1-OE plants were significantly higher than those of WT plants.The distribution in stems and leaves increased the distribution of phosphorus in roots;using soluble phosphorus,cell wall phosphorus,pectin,and pectin methylesterase in shoots and shoots as the criteria for evaluating phosphorus reuse,it was found that LP stress caused Both forms of phosphorus were reduced,but SAMS1-OE reduced cell wall phosphorus and increased soluble phosphorus in plants.Overexpression of SAMS1 increased pectin concentration in both tissues and increased pectin methylesterase activity.In order to further study the absorption and utilization of mineral nutrients by SAMS1during the whole growth period of tomato,we adopted the method of grafting and found that under LP treatment,the aboveground dry weight and fruit yield of SAMS1-OE as rootstock were gradually increased.The ratio of root to shoot was higher than that of WT plants,and the application of phosphorus fertilizer promoted the distribution of phosphorus from branches and roots to fruits.The effect of SAMS1 on the absorption and utilization of phosphorus fertilizer was analyzed by fertilizer utilization rate and yield contribution.The use of transgenic rootstocks increased the amount of phosphorus uptake by plants,and PUE was negatively correlated with the amount of phosphorus application.The contribution of SAMS1to yield gradually decreased with the increase of phosphorus content.The above results show that SAMS1 can promote the degradation of Put and promote the growth of lateral roots;Eth can promote the hydrolysis of the above-ground cell wall phosphorus to soluble phosphorus,increase the root phosphorus concentration,increase the Apase activity in the soil,increase the release of organic phosphorus in the soil,and increase the plant’s ability to absorb phosphorus.Absorption of elements;through field cultivation,it was found that SAMS1 can be grafted as a rootstock,which can improve the utilization rate of phosphorus fertilizer and increase the yield of tomato.It provides theoretical and technical support for improving the utilization efficiency of phosphate fertilizer in practical agricultural applications.更多还原