扶芳藤茎尖的玻璃化法超低温保存及其植株再生

扶芳藤属卫矛科卫矛属常绿藤本灌木,我国拥有丰富的扶芳藤资源,目前已经引起了专家学者的注意,扶芳藤的资源和变异研究(潘青华等2004)、叶片粗蛋白和氨基酸成分分析(潘青华等     

水分胁迫下扶芳藤幼苗保护酶活性和渗透调节物质的变化

扶芳藤(Euonymus fortunei(Turcz.)Hand.-Mazz)作为北方城市园林的新优攀援藤本和地被植物越来越受到重视,新品系的抗逆性筛选工作迫在眉睫。本文采用聚乙二醇(PEG-6000)模拟水分胁迫的方法,测定其生理指标,研究水分胁迫对扶芳藤不同品系幼苗叶片保护酶活性和渗透调节物质的影响,旨在摸索出这些生理指标与植物抗旱性的关系。研究结果表明,随着模拟干旱时间的延长,扶芳藤幼苗叶片中相对含水量(RWC)逐步下降,保护酶活性上升,脯氨酸和可溶性糖含量急剧上升,并且抗旱性强的品系保护酶活性和渗透调节物质含量较高。     

不同油橄榄品种对低温胁迫的生理响应及抗寒性综合评价

为了明确油橄榄的抗寒机理,对甘肃陇南地区主栽的6个油橄榄品种离体叶片进行低温胁迫(5℃、0℃、-5℃、-10℃)处理,并以18℃常温培养为对照,测定与抗寒性相关的生理生化指标,并采用隶属函数法对不同品种抗寒性进行综合评价。结果显示:(1)各油橄榄品种离体叶片相对电导率随处理温度的降低而呈"S"型曲线变化,结合Logistic方程得出不同品种油橄榄半致死温度介于-6.687℃~-3.706℃之间,且各品种的半致死温度顺序为‘皮肖利’<‘配多灵’<‘莱星’<‘阿斯’<‘鄂植8号’<‘佛奥’,其抗寒性大小依次为‘皮肖利’>‘配多灵’>‘莱星’>‘阿斯’>‘鄂植8号’>‘佛奥’。(2)随着胁迫温度降低,各油橄榄品种离体叶片叶绿素含量逐渐降低,可溶性糖含量基本呈逐渐增加趋势,而可溶性蛋白质含量、脯氨酸含量、丙二醛含量、SOD活性、POD活性和CAT活性则呈先升后降的趋势。(3)在-10℃低温胁迫下,油橄榄品种‘皮肖利’和‘配多灵’的叶片叶绿素含量比常温对照降幅较小,渗透调节物质含量和保护酶活性相对增幅较大,丙二醛含量增幅较小,而此时品种‘佛奥’和‘鄂植8号’各指标的变化趋势与之相反。(4)根据不同油橄榄品种的平均隶属度(抗寒性综合评价值)确定的品种抗寒能力大小顺序(‘皮肖利’>‘配多灵’>‘莱星’>‘阿斯’>‘鄂植8号’>‘佛奥’)与品种半致死温度的抗寒性表现顺序完全一致。研究表明,在低温胁迫条件下,‘皮肖利’和‘配多灵’等油橄榄品种能通过提高渗透调节物质含量和增强保护酶活性来缓解胁迫造成的过氧化伤害,从而表现出较强的抗寒能力;基于相对电导率的半致死温度和隶属函数法的综合评价均能准确鉴定油橄榄的抗寒性。     

棉花幼苗对不同程度低温逆境的生理响应

以‘新陆早33号'(冷敏感)和‘中棉所50号’(高耐寒)棉花品种为材料,采用人工模拟低温方法,研究不同温度和时间处理下棉花幼苗叶片抗氧化酶活性和渗透调节物质含量的响应特征,探讨不同温度胁迫下棉花的抗逆机制.结果显示:(1)随胁迫温度降低、胁迫时间延长,两参试棉花品种幼苗遭受低温伤害的程度逐渐加剧,尤以5℃和0℃处理表现最为明显.(2)经15℃及10℃处理后,植株叶片REC、POD活性、SP和Pro含量总体上随胁迫时间延长而提高,MDA含量、APX活性和SS含量则先升后降,并均于处理24 h时达峰值;而在5℃及0℃处理下,REC和SOD、POD活性以及MDA、SS、SP含量迅速上升,APX活性、Pro含量则呈先升后降的变化趋势.(3)与‘新陆早33号,相比,‘中棉所50号’在各处理水平下均可保持更大的抗氧化酶活性和渗透调节物质含量上升幅度.研究表明,在不同程度低温逆境中,棉花幼苗可通过调节抗氧化酶活性和渗透调节物质含量,启动相应抗逆调节方式来维持代谢平衡,增强抗逆性;高耐寒性品种在低温逆境中具有更灵敏、高效的活性氧清除能力和渗透调节响应机制,从而有效抵御低温逆境伤害.     

低温对不同耐寒性橡胶树叶片抗氧化能力的影响

为了探讨低温胁迫对橡胶树耐寒新品种‘湛试327-13’和不耐寒品种‘文昌217’叶片抗氧化能力的影响,以橡胶嫁接苗为材料,模拟8℃/8℃(昼/夜)的低温胁迫,在4d内每天连续测定叶片抗氧化酶活性以及相关的生理参数,再通过激光共聚焦显微镜观察气孔保卫细胞H2O2荧光密度。结果发现:(1)低温胁迫使‘湛试327-13’和‘文昌217’叶片超氧阴离子产生速率、SOD、CAT和POD及APX活性均出现先升后降的趋势,且在整个低温胁迫中,‘湛试327-13’的各抗氧化酶活性均高于‘文昌217’;低温胁迫使‘湛试327-13’的H2O2含量在处理第1天后就达到最大值,‘文昌217’则在第3天才达到最大值。(2)两个品种叶片的H2O2含量和气孔保卫细胞的H2O2荧光强度在低温胁迫后的表现规律与上述指标基本一致。研究表明,‘湛试327-13’叶片超氧阴离子和H2O2快速积累,可能起到能促进抗氧化酶活性的提高,以及降低膜质氧化胁迫程度和TBARS含量的作用;且同时保持了较高的相对含水量和较低的相对电导率,从而提高了其对低温胁迫的耐性。     

盐和水分预处理对扶芳藤幼苗抗寒性的影响

测定了扶芳藤新品种‘红脉’经不同方式预处理后对幼苗抗寒性的影响,测定了膜脂过氧化水平、保护酶活性和渗透调节物质含量的变化。结果表明,与对照低温胁迫相比,0.1和0.3mol/L NaCl溶液预处理后再经低温胁迫处理,扶芳藤幼苗的膜脂过氧化水平较低、保护酶活性高、渗透调节物质含量的累积较多,不同浓度的NaCl溶液预处理比不同浓度的PEG溶液预处理对提高扶芳藤的抗寒性更有效。通过实验证明了植物在不同的逆境中确实存在着交叉适应现象,植物对各种逆境胁迫有着许多相同的生理反应,对各种胁迫的适应性存在某些共同的机制。     

冷驯化提高金边卫矛抗寒力期间细胞ATP酶的超微细胞化学定位

采用磷酸铅沉淀的电镜细胞化学方法,对经冷驯化及未经冷驯化的金边卫矛 EuonymusradicansEmorald&Gold. 以及两者在-5℃下处理1d后的叶肉细胞进行了ATPase的超微细胞化学定位的比较观察,并对冷驯化可提高植物抗寒力及ATPase与植物抗寒性关系进行了探讨.标志ATPase活性反应的磷酸铅沉淀物主要分布在叶片栅栏细胞和海绵细胞的质膜和液泡膜上,ATPase在这两类细胞中的分布及活性无明显差异.金边卫矛幼苗经4℃低温驯化7d后,其质膜和液泡膜的ATPase活性高于22℃下生长的幼苗,冷驯化后的金边卫矛幼苗在-5℃处理1d后,其质膜和液泡膜上仍可观察到ATPase的活性反应,但较低温胁迫前有所降低,且超微结构完整,未见任何伤害.而未经冷驯化的金边卫矛幼苗在-5℃处理1d后,其质膜和液泡膜上的ATPase完全失活,同时细胞的超微结构受到严重伤害.实验结果表明,冷驯化可提高金边卫矛幼苗ATPase在低温胁迫下的稳定性,这种活性变化与植物抗寒力的提高呈正相关,同时还发现液泡膜ATPase活性较质膜ATPase变化幅度大,推测液泡膜可能较质膜ATPase对低温更为敏感.     

2种枸杞叶片对混合盐胁迫的生理响应

选取耐盐性不同的‘宁杞5号’和‘黑杞1号’一年实生苗为试验材料,进行渗透势分别为-0.47、-0.82和-1.18MPa的混合盐(NaCl+Na_2SO_4)胁迫处理,分析比较2种枸杞叶片生理指标的响应特征,以揭示2种枸杞适应盐胁迫的生理机制。结果表明:(1)随着渗透势降低,2种枸杞叶片生理指标响应趋势基本一致,叶绿素(Chl)和可溶性蛋白(SP)含量先升后降,相对电导率、丙二醛(MDA)和脯氨酸(Pro)、可溶性糖(SS)含量逐渐增加,但‘宁杞5号’的变化幅度大于‘黑杞1号’。(2)随着渗透势降低,2种枸杞叶片的超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)活性均增强,但-1.18 MPa混合盐溶液处理则抑制了‘宁杞5号’抗氧化酶活性。(3)随着盐胁迫处理时间的延长,2种枸杞叶片相对电导率、MDA含量、Pro含量、SS含量和CAT活性增加,但‘宁杞5号’的渗透调节物质积累量高于相应‘黑杞1号’;而2种枸杞Chl含量及SOD和POD活性在-1.18 MPa混合盐溶液处理下有所下降。研究表明,2种枸杞均可通过积累渗透调节物质和加强抗氧化酶系统来缓解盐胁迫对自身的伤害,‘宁杞5号’的渗透调节能力较强,而‘黑杞1号’具有更稳定的抗氧化酶系统,从而具有更强耐受盐胁迫的能力。     

外源NO缓解蝴蝶兰低温胁迫伤害的生理机制研究

以蝴蝶兰品种‘台湾黄金’幼苗(苗龄15个月)为试验材料,通过叶面喷施200μmol·L-1的NO供体硝普钠(SNP),低温胁迫(昼/夜:12℃/7℃)处理5d和10d,分别测定叶片电解质渗漏率、丙二醛(MDA)含量、pH、渗透调节物质含量及几种保护酶活性,探讨外源NO缓解蝴蝶兰低温胁迫伤害的生理机制。结果表明:低温胁迫条件下,预施SNP处理可以有效抑制蝴蝶兰叶片电解质渗漏率、MDA含量和pH的上升,显著提高叶片可溶性糖、可溶性蛋白及脯氨酸(Pro)含量,显著延缓超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)等抗氧化酶活性的下降,增强多酚氧化酶(PPO)和苯丙氨酸解氨酶(PAL)的活性。研究认为,外源NO供体SNP可以通过保护蝴蝶兰幼苗的细胞膜系统,增加渗透调节物质含量,提高保护酶活性来减轻低温胁迫对蝴蝶兰幼苗的伤害,提高其抗低温胁迫的能力。     

PEG胁迫及复水对不同抗旱性小麦幼苗脯氨酸代谢关键酶活性的影响

以两种不同抗旱性小麦品种幼苗为试验材料,采用PEG模拟干旱胁迫处理,探究干旱胁迫及复水对小麦幼苗叶片与根系脯氨酸累积及关键酶活性的影响。结果显示:(1)PEG胁迫下抗旱品种‘普冰143’根长和根干重下降不大,而水敏感品种‘郑引1号’根长和根干重下降显著;且于胁迫处理36h时‘普冰143’根系脯氨酸含量增加(75.0%)显著大于‘郑引1号’(37.7%),复水24h后均恢复至对照水平。(2)PEG胁迫下‘普冰143’叶片中谷氨酸合成途径关键酶P5CS和鸟氨酸合成途径关键酶δ-OAT活性均显著增加,且‘普冰143’叶片脯氨酸两条合成途径关键酶活性均得以加强;PEG胁迫处理36h时,‘郑引1号’叶片中P5CS活性增加显著,δ-OAT活性变化较小,且‘郑引1号’叶片脯氨酸合成可能以谷氨酸途径为主;但在PEG胁迫下两个不同抗旱性品种的根中P5CS、δ-OAT活性均变化较小。(3)PEG胁迫处理36h时‘普冰143’叶片脯氨酸降解酶PDH活性显著下降,而‘郑引1号’叶片PDH活性显著增加,复水后抗旱品种叶片该酶活性显著增加,水敏感品种恢复至对照水平;但PEG胁迫处理下两个不同抗旱性品种的根中PDH活性均显著下降。研究表明,PEG胁迫下小麦叶片是合成脯氨酸的主要部位,抗旱品种‘普冰143’根系脯氨酸持续积累与叶片中高的脯氨酸合成关键酶活性及脯氨酸转运有关。     

Cold shock response inLactococcus lactis ssp.diacetylactis

The acquired freeze-thaw tolerance was investigated forLactococcus lactis ssp.diacetylactis. Pretreatment of microorganisms at less severe temperatures to initiate cold tolerance gaveL. lactis ssp.diacetylactis improved cell viability after successive freezings and thawings. The ability of cells to survive freeze-thaw was dependent on factors experienced prior to freezing. Factors affecting lactic acid bacteria survival during freeze-thaw cycles were found to be different diluents, growth phase, and different cold temperatures. Viability experiments showed that this strain displaying cold shock cryotolerance had an improved survival capacity in stationary phase. The plasmid contents of lactic acid bacteria isolated from different types, DRC-2 and DRC-2C, were examined and compared with the plasmid contents of culture collection strains both before and after cold shock treatment. Using agarose gel electrophoresis, no obvious correlation between the cold shock response and the number of plasmids in the cell could be observed.     

天山雪莲SiICE2基因克隆及抗寒性分析

该研究以天山雪莲(Saussurea involucrata)的转录组数据为基础,利用Premier 5.0设计1对特异性引物SiICE2-Up和SiICE2-Down,以天山雪莲cDNA为模板克隆得到天山雪莲SiICE2基因的开放阅读框(ORF),对其进行生物信息学分析;构建植物表达载体pCAMBIA2300-35S-SiICE2-Nos,利用农杆菌介导法导入番茄(Lycopersicon esculentum),通过PCR和RT-PCR对转化植株进行验证,qRT-PCR分析转SiICE2基因番茄株系SiICE2基因的相对表达量;将转SiICE2基因型和野生型番茄在0℃处理后,进行抗寒性分析。结果表明:(1)成功克隆得到天山雪莲SiICE2基因,其大小为462bp,共编码153个氨基酸,系统进化分析发现SiICE2蛋白与菜蓟(Cynara scolymus L)亲缘关系最近。(2)成功构建了植物表达载体pCAMBIA2300-35S-SiICE2-Nos,经农杆菌介导法侵染番茄,PCR鉴定表明共有9株为转SiICE2基因番茄植株。(3)膜生理指标测定结果显示,随着低温处理时间的增加,转SiICE2基因型番茄的相对电导率、丙二醛含量均显著低于野生型,在处理时间为24h时,转SiICE2基因型番茄相对电导率比野生型低31.7%,丙二醛含量比野生型低4.2μmol/g。(4)抗氧化酶活性测定结果显示,随着低温处理时间的增加,转SiICE2基因番茄植株的POD、CAT和SOD活性均呈现持续递增趋势,野生型呈先逐渐升高后降低的趋势,且各处理时间内转SiICE2基因番茄的POD、CAT和SOD活性均显著高于野生型。研究发现,天山雪莲SiICE2基因可以显著增强非低温驯化番茄的抗寒性。     

Cold acclimation and cold-regulated gene expression in ABA mutants of Arabidopsis thaliana

We have examined the cold-induced enhancement of freezing tolerance and expression of cold-regulated (cor) genes in Arabidopsis thaliana (L.) Heynh (Landsberg erecta) and abscisic acid (ABA)-deficient (aba) and ABA-insensitive (abi) mutants derived from it. The results indicate that the abi mutations had no apparent effect on freezing tolerance, while the aba mutations did: cold-acclimated aba mutants were markedly impaired in freezing tolerance compared to wild-type plants. In addition, it was observed that non-frozen leaves from both control and cold-treated aba mutant plants were more ion-leaky than those from corresponding wild-type plants. These data are consistent with previous observations indicating that ABA levels can affect freezing tolerance. Whether ABA has a direct role in the enhancement of freezing tolerance that occurs during cold acclimation, however, is uncertain. Several studies have suggested that ABA might mediate certain changes in gene expression that occur during cold acclimation. Our data indicate that the ABA-induced expression of three ABA-regulated Arabidopsis cor genes was unaffected in the abi2, abi3, and aba-1 mutants, but was dramatically impaired in the abi1 mutant. Cold-regulated expression of all three cor genes, however, was nearly the same in wild-type and abi1 mutant plants. These data suggest that the cold-regulated and ABA-regulated expression of the three cor genes may be mediated through independent control mechanisms.     

Cold acclimation and photoinhibition of photosynthesis in Scots pine

Cold acclimation of Scots pine did not affect the susceptibility of photosynthesis to photoinhibition. Cold acclimation did however cause a suppression of the rate of CO₂ uptake, and at given light and temperature conditions a larger fraction of the photosystem II reaction centres were closed in cold-acclimated than in nonacclimated pine. Therefore, when assayed at the level of photosystem II reaction centres, i.e. in relation to the degree of photosystem closure, cold acclimation caused a significant increase in resistance to photoinhibition; at given levels of photosystem II closure the resistance to photoinhibition was higher after cold acclimation. This was particularly evident in measurements at 20° C. The amounts and activities of the majority of analyzed active oxygen scavengers were higher after cold acclimation. We suggest that this increase in protective enzymes and compounds, particularly Superoxide dismutase, ascorbate peroxidase, glutathione reductase and ascorbate of the chloroplasts, enables Scots pine to avoid excessive photoinhibition of photosynthesis despite partial suppression of photosynthesis upon cold acclimation. An increased capacity for light-induced de-epoxidation of violaxanthin to zeaxanthin upon cold acclimation may also be of significance.Abbreviations APX ascorbate peroxidase - DHA dehydroascorbate - DHAR dehydroascorbate reductase - Fm maximal fluorescence when all reaction centres are closed - Fv/Fm maximum photochemical yield of PSII - GR glutathione reductase - GSH reduced glutathione - Je rate of photosynthetic electron transport - MDAR monodehydroascorbate reductase - q_N nonphotochemical quenching of fluorescence - q_P photochemical quenching of fluorescence - SOD superoxide dismutase This work was supported by the Swedish Natural Science Research Council and the National Natural Science Foundation of China.     

五种金花茶组植物的耐寒性比较研究

金花茶组植物是世界珍稀、濒危的观赏植物,具有极高的观赏价值和药用价值.为了比较金花茶组植物的耐寒性,以五种金花茶组植物为材料,采用人工模拟低温环境的方法对其2年生叶片进行低温胁迫处理,应用电导法研究五种金花茶种质在20℃(常温对照)、8℃、-2℃、-7℃、-12℃、-17℃、-22℃和-27℃低温下相对电导率的变化,配合Logistic方程,测定其低温半致死温度(LT50),以及叶片中游离脯氨酸、可溶性糖和丙二醛的含量.结果表明:五种金花茶的低温半致死温度(LT50)范围为-14.58~-12.74℃,其中金花茶为-14.58℃、龙州金花茶为-14.27℃、柠檬黄金花茶为-13.44℃、直脉金花茶为-13.09℃、东兴金花茶-12.74℃.低温半致死温度能反映金花茶种质的耐寒性,金花茶和龙州金花茶耐寒性强,其次为柠檬黄金花茶和直脉金花茶,东兴金花茶耐寒性较弱.在降温过程中,五种金花茶叶片相对电导率随温度降低呈S型上升,与温度呈负相关;脯氨酸、可溶性糖和丙二醛含量均呈现先上升后下降的趋势.同一低温条件下,半致死温度低的金花茶脯氨酸和可溶性糖的含量更高,而丙二醛含量更低.该研究结果为金花茶组植物耐寒种质选育提供了科学依据,为人工种植金花茶提供了技术支持.     

Cold stored ectoparasitoid of Cydia fruit moths released under different temperature regimes

Cold storage of parasitoids to be used in biological control programs is desirable but risky for the performance of the stored parental generation as well as for its offspring. We studied the performance of cold stored and unstored parasitoids after release at different temperature regimes in the laboratory at the level of two subsequent generations in Hyssopus pallidus (Askew) (Hymenoptera: Eulophidae). This gregarious ectoparasitoid is a candidate biocontrol agent of Cydia pomonella L. (Lepidoptera: Tortricidae) and Cydia molesta (Busck) (Lepidoptera: Tortricidae) larvae, two fruit pests of high economic significance in apple cultivation. Cold storage for 14 days at 4°C imposed to the pupal stage of the parasitoid did not reduce the parasitism capacity of the parental generation, nor did it alter the female biased sex ratio of the progeny. Remarkably, this short-term storage of the parental generation exhibited a significant and consistently positive effect on offspring weight throughout all ambient temperature regimes, resulting in an increased offspring weight. Furthermore, offspring number was only reduced after release at low ambient temperatures, but not at 25°C and 30°C. Irrespective of whether the parasitoids originated from the stored or unstored group, highest parasitism rate was achieved at temperatures above 20°C. In conclusion, this candidate biocontrol agent can be cold stored for short periods without any measurable quality loss after release at most except at low ambient temperatures. Our findings suggest that H. pallidus is a thermophilous parasitoid that will perform best when applied at warm ambient temperatures in fruit orchards.     

Cold pretreatment enhances microspore embryogenesis in oilseed rape (Brassica napus L.)

Stress is an essential component during embryogenesis induction in microspore culture. Cold pretreatment has been used in cereal microspore culture but very seldom attempted in Brassica microspore culture. The effect of cold pretreatment of flower buds subjected to a liquid medium on microspore embryogenesis was investigated in spring and winter Brassica napus, as well as in B. rapa and B. oleracea. Cold pretreatment significantly enhanced microspore embryogenesis (by 1–7 fold) compared to commonly used microspore culture protocol in B. napus, while it was less effective in B. rapa or even negative in B. oleracea. The appropriate duration of cold pretreatment was found to be 2–4 days, which stimulated the best microspore embryogenesis. Cold pretreatment was also able to promote embryo development including the improvement of embryo quality and acceleration of embryogenesis. When incorporating with medium refreshing, cold pretreatment could initiate the most microspore embryogenesis than any other treatment used. With further improvement cold pretreatment method may have a positive potential in Brassica breeding programmes.     

Comparison of Barley Response to Short-Term Cold or Dehydration

Abscisic acid (ABA) content and relative water content (RWC) in second fully expanded leaves of cold hardened plants and in dehydrated leaves of freezing tolerant barley (Hordeum vulgare L. cv. Lunet) were compared. ABA content and RWC in leaves did not change during the first day of cold hardening. On the contrary, dehydration of leaves led to a decrease of RWC and to an increase of ABA content.     

Cold storage of Trichogramma ostriniae reared on Sitotroga cerealella eggs

Efficient storage of the biological controlagent Trichogramma ostriniae couldimprove current parasitoid production methodsby making the system more flexible andefficient. Initial studies compared emergencerates of T. ostriniae reared onSitotroga cerealella eggs held at 6 °C, 9 °C, 12 °C,15 °C, and 24 °C for up to 8 weeks after parasitism.At 15 °C, emergence occurred in <2 weeks.Emeregence was >80% for parasitized eggsstored at 9 °C and 12 °C for 4 and 6 weeksrespectively. Storage at 6 °C caused asignificant decline in emergence after 2 weeks. Subsequent trials focused on fitness of storedT. ostriniae. Percentage of emergedfemales parasitizing eggs, female longevity,and fecundity were quantified after storage.The percentage of females successfullyparasitizing O. nubilalis eggs wasgreatest for the 9 °C four-week treatment(100%). Compared to 24 °C controls, storage at12 °C for 6 weeks or at 9 °C for 8 weeks reduced thepercentage of females parasitizing. Longevityof females held in cold storage was generallyless than that of controls. Rates of parasitismby stored Trichogramma was generallysimilar to controls after 2 to 4 weeks' storageat 9 °C and 12 °C but declined with storage longerthan 4 weeks. Emergence of progeny of cold-storedfemales was lower than controls for alltreatments. The percentage of female progenyfrom cold stored females was comparable tocontrols up to 4 weeks of storage.