【作者】 李益清；

【导师】 李天来；

【作者基本信息】 沈阳农业大学 ， 蔬菜学， 2011， 博士

【摘要】 番茄是我国设施栽培的重要果菜之一,然而在设施栽培过程中经常会遭遇弱光逆境,使其光合生产力的发挥受到了抑制,并最终导致产量降低,商品性变差,给生产造成严重损失。因此,研究番茄在弱光逆境下的生理生化变化及其机制,对设施番茄高产优质栽培及耐弱光品种选育鉴定具有关键作用。尽管之前关于番茄弱光的报道较多,但多是结合低温进行的,并未将其作为单一因素进行研究。本试验以栽培型番茄W(L. esculentum.Mill)为试材,通过对其进行弱光和弱光下的钙素调控处理,研究了短期弱光及其恢复对番茄幼苗生长发育、光合作用、光合电子传递、活性氧代谢等方面的影响。此外,还研究了外源钙素对弱光下番茄叶片解剖结构和细胞超微结构、叶绿素合成、光合特性、光合电子传递等方面的调控作用,以期进一步了解弱光对番茄光合生理代谢的具体作用机制和钙素调控机制,从而为设施番茄栽培和化学物质调控以及抗逆生理育种提供理论依据。主要研究结果如下：1.明确了短期弱光促进番茄幼苗伸长生长,抑制茎的增粗生长,减少了叶片干物质积累,叶绿素含量先升后降,净光合速率的降低主要由非气孔限制因素引起,且羧化效率及RuBP最大再生速率降低。6d以内弱光处理后恢复相应天数,番茄株高、叶绿素含量可基本恢复到对照水平,而茎粗、干物重、和各项光合指标则需要将近2倍处理天数才可恢复到对照水平；9d以上弱光处理后番茄幼苗株高、茎粗和叶片干重、Pn、Gs、Ci、Ls和羧化效率及RuBP最大再生速率则均未能在短期内恢复到对照水平。2.明确了短期弱光降低了番茄幼苗叶片的PSII的光化学活性和电子传递速率,增加了热耗散系数。6d以内弱光处理各项指标与对照之间差异不显著,恢复相应天数后即接近对照水平；9d以上弱光处理显著降低了番茄幼苗叶片的Fv/Fm、Fv/Fo、1/Fo-1/Fm、Y(Ⅱ)、ETR(Ⅱ)及qP和qN值,增加了Fo,自然光恢复阶段虽有所回升但很难恢复到对照植株水平,说明对PSII造成了不可逆转的伤害。3.明确了短期弱光增加了番茄幼苗叶片内的活性氧含量,降低了保护酶活性及可溶性蛋白含量。处理初期保护酶活性应激性增加的同时活性氧含量下降,6d以上弱光处理则显著增加了活性氧水平,降低了保护酶活性,叶片膜脂过氧化程度提高,但在后期自然光条件下可基本达到对照植株水平,而9d以上处理则很难在短期内恢复到对照水平。短期弱光处理对POD活性的影响较小。4.明确了外源钙素的加入提高了弱光下番茄植株的高度,增加了茎粗,均衡了植株地上和地下部分的生长；同时也增加了植株各部分干物质积累量,使得向根系和果实分配的干物质比例增加,亦均衡了植株营养生长和生殖生长；钙素的加入增加了叶片中的Chl含量,提高了植株光合能力,增加了同化物的生产力,增强了保护酶活性,从而提高了弱光下番茄的产量和品质。IAA可在一定程度上放大钙素的调控效果。5.明确了番茄幼苗在弱光处理后,叶片厚度、栅栏组织和海绵组织厚度均较自然光对照发生不同程度下降,气孔器显著增大,气孔开口程度和开口率也有所增加,气孔数极显著增大,栅栏组织大小极显著降低,而栅栏组织内叶绿体数目有所增加,叶绿体数目显著增加,单个叶绿体和叶绿体内的淀粉粒大小以及淀粉粒、嗜饿颗粒和基粒数目均显著下降。钙素显著提高了弱光条件下番茄幼苗的叶片厚度和栅栏组织厚度,海绵组织厚度也较弱光对照有所增加,但作用不显著；显著增加了气孔器大小和开口程度,降低了气孔张开率和气孔数；极显著增加了栅栏组织大小和栅栏组织内的叶绿体数目,甚至极显著的高于自然光对照水平；钙素的加入也显著增加了叶绿体数目和单个叶绿体大小及叶绿体内的淀粉粒数、嗜锇颗粒数以及基粒数,在基粒数方面较自然光对照提高5.44%,但二者间差异不显著。6.明确了弱光处理后番茄幼苗叶片中叶绿素合成前体PBG含量较自然光对照植株显著下降,而UrogenⅢ和CorprogenⅢ含量则较自然光对照植株增加,ProtoⅨ、Mg-ProtoⅨ和Pchl含量变化规律不太明显,说明弱光下叶绿素合成受阻位点在PBG、UrogenⅢ和CorprogenⅢ的转化过程中；弱光处理后番茄叶片中的Chlase活性显著增加,说明其对叶绿素的降解增强；且Chl含量呈先增后降的变化趋势,Chla/b比值下降。外源钙素的加入显著提高了弱光下番茄叶片Chl合成途径中的各前体物质含量,较弱光对照降低了Chlase酶活,减缓了Chl的酶降解过程,增加了弱光下番茄叶片中Chl含量,对Chla/b的影响不是很明显。7.明确了弱光处理后番茄幼苗叶片Pn、Gs和Tr值显著下降,Ci值增加,PSI的P700还原态水平和PSII光化学活性降低,两个光系统的量子产额和电子传递速率下降,分配给光反应的能量比例降低,热耗散加强,光系统间能量分配的不平衡性增大,光合作用受抑,且PQ库容积减小,引发了围绕PSI的环式电子流。钙素通过增加Gs和Tr值,降低Ci值提高了Pn值；钙素处理也提高了P700还原态水平,增强了光系统的光化学活性,使向光反应部分分配的光能比例增加,增强了热耗散系数,均衡了光系统间的能量分配比例,扩大了PQ库容量,加速了围绕PSI的环式电子流传递速度。8.明确了弱光处理使番茄叶片内Rubisco初始活力、总活力和活化度在处理3d时应激性增加,之后随处理时间的延长逐渐下降；弱光处理降低了番茄叶片内与光合碳同化有关的酶活性,抑制了cab、rca和rbc S基因的相对表达量。外源钙素的加入提高了弱光下番茄叶片中Rubisco的初始活力、总活力及活化程度,PEPCase、FBPase和GO活性也较弱光对照增强,显著提高了弱光下番茄幼苗叶片中cab、rca和rbc S三个基因的相对表达量,提高了叶片的同化物生产及运输能力。9.明确了弱光处理后番茄幼苗叶片内活性氧水平在处理初期下降,随着处理时间的延长逐渐增加,叶片MDA和LOX含量在弱光处理后显著降低；抗氧化酶活性的变化趋势与活性氧水平相反,处理3d时迅速增加,之后逐渐下降；GSH和AsA含量的变化趋势类似于抗氧化酶活性变化趋势,处理3d时增加,之后逐渐下降；Pro含量则始终显著高于自然光对照水平,而可溶性糖和可溶性蛋白含量在处理6d以后显著低于自然光对照。钙素处理在一定程度上降低了弱光胁迫下番茄幼苗叶片内的活性氧水平,减轻了膜脂过氧化程度,提高了各抗氧化酶活性,减轻了活性氧对植株造成的伤害,且钙素处理的番茄幼苗叶片内各渗透调节物质含量均高于弱光对照水平。更多还原

【Abstract】 Tomato is one of the important vegetables in greenhouse cultivation, but it often encountered low-light stress during cultivation, photosynthetic productivity was inhibited, and eventually lead to lower production and quality, causing serious losses to the production. Therefore, the study of tomato under low light stress and its physiological and biochemical mechanisms of change, high yield and quality of facilities tomato cultivation and breeding resistance to low-light identification plays a key role. Although previous reports on the more low light tomato, but mostly carried out with cold, not as a single factor study. In this experiment, cultivation of tomato W (L.esculentum.Mill) as material, through its low-light treatment and control measures of calcium under low light to study the short-term light stress and its recovery on the tomato seedling growth, photosynthetic the role of electron transfer, reactive oxygen metabolism, in addition of exogenous calcium to its anatomical structure and cell ultrastructure, chlorophyll synthesis, photosynthesis, photosynthetic electron transport regulation of other aspects to further understanding of the weak Light on Photosynthetic physiology of the specific mechanism of action and mechanism of calcium regulation, so as to chemical substances for greenhouse cultivation and breeding of stress physiological regulation and provide a theoretical basis. The main results are as follows:1. Through short-term low-light and the recovery treatment, we found that plant height increased, stem diameter decreased, dry matter weight in leaves was decreased, the chlorophyll content of tomato leaves first increased and then decreased, Pn reduce mainly caused by non-stomatal limitation, and CE and Jmax values decreased.6 days or less low light treatment and natural light recovery the same days, plant height, chlorophyll content can be achieved to contrl level, but stem diameter, dry matter weight and photosynthesis parameters need about 2 times the processing time required to recover; after 9 days or more low light treatment, plant hight, stem diameter, leaf dry weight, Pn, Gs, Ci, Ls, CE and Jmax value were more difficult to restore.2. Clear short-term low intensity light treatment decreased the photo-chemical activity and electron transmit rate, increased heat dissipation coefficient. Within 6 days low light treatment, above parameters have no significant difference between treatment and control, the same time recovery after treatment can close to the control level; 9 days or more treatment can redueced Fv/Fm, Fv/Fo,1/Fo-1/Fm, Y (Ⅱ), ETR (Ⅱ), qP and qN, enhanced Fo, although there was found a pick-up in natural lilght recovery condition, but can’t completely restored to control levels, which declare that low light intensity treatment cause irreversible damage to PSⅡ.3. Low light and its recovery through short-term treatment, enhanced active oxyzen contents, reduced protected enzymes activities and the content of soluble protein. The activities of protected enzymes increased in dealing with the initial stress, but the content of active oxyzen decresed, above 6 days treatment, the active oxyzen level was increased significantly, protected enzymes activities decresed, and the degree of membrane little influence on lipid peroxidation enhanced, but they can be restored after a certain number of days to reach the basic level of control,9 days or more was difficult to return to the control level. There was little influence of low light intensity treatment to the activity of POD.4. Through the first flowering when the first inflorescence of tomato plants with low light low light treatment and exogenous calcium and IAA spraying treatment, defined calcium enhanced plant height and stem diameter of tomato plant, balanced above and below ground parts of the plant growth; increased the dry matter weight in various parts of tomato plants, and increased dry matter allocation to the roots and the proportion of fruit, balance the vegetative and reproductive growth; The addition of calcium were increased tomato leaf Ch1 content and Pn, Gs and Tr, lower Ci, that they participate in tomato leaves under low light intensity regulation of stomatal movement; clear exogenous calcium can be increase under low light tomato leaf and fruit of glucose, fructose and sucrose content; clear calcium has a positive regulatory role in leaves of tomato plants under low light defensive activity; Calcium can be increased the protected enzymes activities, soluble protein content, photosynthesis, membrane lipid peroxidation to some extent in tomato leaves under low light rnvironment, and reduce the MDA content, but POD activity in the regulation of calcium in terms of promotion is very limited. In addition, calcium can increase the tomato fruit weight, yield numbers of fruits to enhance the fruit soluble solids, soluble sugars, organic acids and Vc content and sugar acid ratio, and improve the fruit quality under low light conditions. IAA in the above areas could enlarge the role of calcium regulation.5. Through the leaves as one of six low-light tomato seedlings under low light stress treatment and calcium handling, clear low-light treatment of tomato leaf thickness, palisade tissue and spongy tissue thickness compared with control plants, natural light, had decreased to varying degrees, calcium hormone significantly increased leaf thickness and palisade tissue thickness, spongy tissue thickness increased less light control, but the effect was not significant; low light treatment significantly increased the size of stomata, stomatal opening and the opening degree of the increase in rate, stomatal number of very significant increase of calcium treatment significantly increased the degree of stomatal size and openings, reducing the number of stomata and stomatal opening rate; low light treatment, the size of palisade tissue was significantly reduced, while the palisade tissue increase in the number of chloroplast, calcium treatment significantly increased the palisade tissue size and palisade tissue within the chloroplast number, or even significantly higher than natural light control level; low light treatment, the number of chloroplast increased significantly, a single chloroplast and chloroplast starch grain size and starch grains, hungry, addicted to the number of particles and matrix grains were significantly decreased, calcium significantly increased chloroplast number and chloroplast size and chloroplast single starch grains, osmiophilic particles and the matrix grains, in terms of basic grains was increased by a more natural light 5.44%, but the difference was not significant between them.6. Through the leaves as one of the six low-light treatment of tomato seedlings under low light stress and calcium handling, a clear tomato leaves after a low light the chlorophyll precursor ALA, PBG content in natural light than the control plants decreased significantly, while the UrogenⅢand CorprogenⅢcontent of control plants increased more natural light, ProtoⅨ, Mg-ProtoⅨand Pch1 content variation is not obvious, that the chlorophyll synthesis under low light stress blocked sites in the PBG, UrogenⅢand CorprogenⅢthe transformation process, the addition of exogenous calcium significantly increased the biosynthesis of the Chi precursors content; tomato leaves under low light stress in a more natural light control Chlase activity increased significantly, indicating that increased degradation of chlorophyll, and calcium handling weak light was reduced by enzyme activity, slowing the process of Chi degradation enzyme; light stress in leaves of tomato after the Chla, b and total Chi content was first increased and then decreased trend, Chla/b decreased calcium treatment increased the tomato under low light stress Chi content in leaves of Chla/ b of the impact is not obvious.7. Through the leaves as one of the six low-light treatment of tomato seedlings under low light stress and calcium handling, a clear tomato leaves after a low light value of Pn, Gs and Tr values were significantly decreased, Ci values increased by increasing calcium Gs values and Tr values, reduce the value of Ci is increased by the Pn; clear the low light treatment reduced the level of PSI in the P7oo reduction state, an increase of PSⅡof Fo and Fm, decreased Fv/ Fm, Fv/Fo and 1/Fo-1/Fm equivalents, an increase of calcium handling of the P700 reduction state level of PSⅠ, PSⅡreduced the Fo and Fm, but also increased the Fv/Fm, Fv/Fo and 1/Fo-1/Fm value; weak PSⅠlight stress reduces the Y (Ⅰ) and ETR (Ⅰ) value, an increase of Y (ND) and Y (NA) value of calcium treatment increased the Y (Ⅰ) and ETR (Ⅰ) value, reducing the Y (ND) and Y (NA) value; low light treatment to reduce the PSⅡof the Y (Ⅱ) and ETR (Ⅱ) value, an increase of Y (NPQ) and Y (NO) value of calcium treatment increased the Y (Ⅱ) and ETR (Ⅱ) value, reducing Y (NPQ) and Y (NO) value; low light treatment increased qN values, Ex Hd values, and values and (A+Z)/(V+A+Z) ratio, reduced qL and Pc values of calcium treatment increased the qL and Pc values lower qN values, Ex Hd value and values and (A+Z)/(V+A+Z) ratio; In addition, the low light treatment increased the L (PFD) andβ/α-1 value of calcium treatment reduced the values of more than two; low-light treatment reduces the size of the PQ pool, causing the ring around the PSI electron flow, the addition of calcium increased under low light the PQ pool size, increased the ring around the PSI electron flow transfer speed.8. Through the leaves as one of the six low-light treatment of tomato seedlings under low light stress and calcium handling, clear low light tomato leaves after the initial Rubisco activity, total activity and activation degree of stress when dealing with an increased 3d then gradually decreased with treatment time, the addition of exogenous calcium increased the initial Rubisco activity, total activity and activation; low light treatment to reduce the tomato leaves of PEPCase, FBPase, and GO activity, calcium handling plant leaves of PEPCase, FBPase, and GO activity of weak optical control enhancements; In addition, low light treatment, the tomato seedlings in the cab, rca and rbc S gene relative expression level of more natural light were decreased significantly, calcium treatment significantly increased the weak Under light stress in leaves of tomato seedlings over the relative expression of three genes.9. Through the leaves as one of the six low-light treatment of tomato seedlings under low light stress and calcium handling, clear low-light treatment of tomato leaves O2·- production rate, H2O2 content and·OH is lower than the initial content of natural light in the treatment the level of control after treatment time with the gradual increase in leaf MDA content and LOX significantly reduced low-light treatment; calcium treatment to some extent reduce the leaves of tomato seedlings under low light within the level of reactive oxygen species, reducing lipid peroxidation; clear low light tomato leaves after antioxidant enzymes SOD, POD, CAT and APX activity in the opposite trend and the level of reactive oxygen species, when dealing with the rapid increase in 3d, then decreased gradually; calcium treatment increased The activities of antioxidant enzymes, thereby reducing the reactive oxygen species damage to the plants; clear low light tomato leaves treated with GSH and AsA content in the trend of changes in antioxidant enzyme activities similar to trends,3d, increase, then decreased gradually, Pro content in leaves were always significantly higher than control level of natural light, while the soluble sugar and soluble protein content after treatment was significantly lower than 6d natural light control; calcium handling within the tomato seedlings were higher than the osmotic adjustment of low-light level of the control content.更多还原