Evaluation of Genetic and Epigenetic Modification in Rapeseed （Brassica napus） Induced by Salt Stress

Salinity is an important limiting environmental factor for rapeseed production worldwide. In this study, we assessed the extent and pattern of DNA damages caused by salt stress in rapeseed plants. Amplified fragment length polymorphism （AFLP） analysis revealed dose-related increases in sequence alterations in plantlets exposed to 10-1000 mmol/L sodium chloride. In addition, individual plantlets exposed to the same salt concentration showed different AFLP and selected region amplified polymorphism banding patterns. These observations suggested that DNA mutation in response to salt stress was random in the genome and the effect was dose-dependant. DNA methylation changes in response to salt stress were also evaluated by methylation sensitive amplified polymorphism （MSAP）. Three types of MSAP bands were recovered. Type Ⅰ bands were observed with both isoschizomers Hpa Ⅱ and Msp Ⅰ, while type Ⅱ and type Ⅲ bands were observed only with Hpa Ⅱ and Msp Ⅰ, respectively. Extensive changes in types of MSAP bands after NaCI treatments were observed, including appearance and disappearance of type Ⅰ, Ⅱ and Ⅲ bands, as well as exchanges between either type Ⅰand type Ⅱ or type Ⅰ and type Ⅲ bands. An increase of 0.2-17.6% cytosine methylated CCGG sites were detected in plantlets exposed to 10- 200 mmol/L salt compared to the control, and these changes included both de novo methylation and demethylation events. Nine methylation related fragments were also recovered and sequenced, and one sharing a high sequence homology with the ethylene responsive element binding factor was identified. These results demonstrated clear DNA genetic and epigenetic alterations in planUets as a response to salt stress, and these changes may suggest a mechanism for plants adaptation under salt stress.     

Seed Treatment with Auxins Modulates Growth and Ion Partitioning in Salt-stressed Wheat Plants

Experiments were performed to determine whether seed priming with different concentrations （100, 150, and 200 mg/L） of auxins （indoleacetic acid （IAA）, indolebutyric acid （IBA）, or their precursor tryptophane （Trp）） could alter salinity induced perturbances in salicylic acid and ion concentrations and, hence, growth in wheat （Triticum aestivum L.） cultivars, namely M.H.-97 （salt intolerant） and tnqtab-91 （salt tolerant）. Primed and non-primed seeds were sown in Petri dishes in a growth room, as well as in a field treated with 15 dS/m NaCl salinity. All priming agents, except IBA, increased the final germination percentage in both cultivars. The seedlings of either cultivar raised from Trp-treated seeds had greater dry biomass when under salt stress. In field experiments, Trp priming was much more effective in mediating the increase in grain yield, irrespective of the cultivar, under salt stress. The alleviatory effect of Trp was found to be associated with reduced uptake of Na^＋ in the roots and subsequent translocation to the shoots, as well as increased partitioning of Ca^＋ in the roots of salt-stressed wheat plants. Plants of both cultivars raised from Trp-and IAA-treated seeds accumulated free salicylic acid in their leaves when under salt stress. Overall, the Trp priming-induced improvement in germination and the subsequent growth of wheat plants could be related to ion homeostasis when under salt stress. The possible involvement of salicylic acid in the Trp priming-induced better growth under Conditions of salt stress is discussed.     

酵母耐盐机制的研究进展

酵母是一种真核模式生物同时也是一种耐盐微生物,其基因表达和信号传导系统的调节机制及离子运输机制与高等真核生物类似。酵母耐盐机制的研究有助于阐明真核生物的耐盐机制。本文综述了酵母在盐胁迫下的信号传导途径和分子应答机制,以及在酵母耐盐机制研究中主要的研究方法。 Abstract:Yeast is a model eukoryotic organism and salt-tolerant microorganism.The regulative mechanism of gene expression and signal transduction and ion transport of yeast is similar to that of higher eukoryotic organism.The research on salt-tolerant mechanism of yeast will be helpful to the illustrate the salt-tolerant mechanism of higher eukoryotic organism.This review summarized the signal transduction pathway and molercular responses of yeast under salt stress and the major research methods in the research on the salt-tolerant mechenism in yeast.     

Seed Treatment with Auxins Modulates Growth and Ion Partitioning in Salt-stressed Wheat Plants

Experiments were performed to determine whether seed priming with different concentrations (100, 150, and 200 mg/L) ofauxins (indoleacetic acid (IAA), indolebutyric acid (IBA), or their precursor tryptophane (Trp)) could alter salinity inducedperturbances in salicylic acid and ion concentrations and, hence, growth in wheat (Triticum aestivum L.) cultivars, namelyM. H.-97 (salt intolerant) and Inqlab-91 (salt tolerant). Primed and non-primed seeds were sown in Petri dishes in a growthroom, as well as in a field treated with 15 dS/m NaCl salinity. All priming agents, except IBA, increased the final germinationpercentage in both cultivars. The seedlings of either cultivar raised from Trp-treated seeds had greater dry biomass whenunder salt stress. In field experiments, Trp priming was much more effective in mediating the increase in grain yield,irrespective of the cultivar, under salt stress. The alleviatory effect of Trp was found to be associated with reduced uptakeof Na~ in the roots and subsequent translocation to the shoots, as well as increased partitioning of Ca~(2 )in the roots ofsalt-stressed wheat plants. Plants of both cultivars raised from Trp-and IAA-treated seeds accumulated free salicylic acidin their leaves when under salt stress. Overall, the Trp priming-induced improvement in germination and the subsequentgrowth of wheat plants could be related to ion homeostasis when under salt stress. The possible involvement of salicylicacid in the Trp priming-induced better growth under conditions of salt stress is discussed.     

Overexpression of a tobacco small G protein gene NtRop1 causes salt sensitivity and hydrogen peroxide production in transgenic plants

The small GTPases of Rop/Rho family is central regulators of important cellular processes in plants. Tobacco small G protein gene NtRop1 has been isolated; however, its roles in stress responses were unknown. In the present study, the genomic sequence of NtRop1 was cloned, which has seven exons and six introns, similar to the Rop gene structure from Arabidopsis. The NtRop1 gene was constitutively expressed in the different organs whereas the other six Rop genes from tobacco had differential expression patterns. The expression of the NtRop1 gene was moderately induced by methyl viologen, NaCl, and ACC treatments, but slightly inhibited by ABA treatment, with no significant induction by NAA treatment. The transgenic Arabidopsis plants overexpressing the NtRop1 showed increased salt sensitivity as can be seen from the reduced root growth and elevated relative electrolyte leakage. The hydrogen peroxide production was also promoted in the NtRop1-trangenic plants in comparison with wild type plants. These results imply that the NtRop1 may confer salt sensitivity through activation of H2O2 production during plant response to salt stress.     

水杨酸和阿斯匹林对小麦幼苗生长过程中盐害的缓解作用

以小麦为材料,研究盐分胁迫对小麦幼苗生长的影响以及水杨酸和阿斯匹林对小麦幼苗生长过程中盐害的缓解作用。结果表明,水杨酸和阿斯匹林能够相对提高盐分胁迫条件下小麦幼苗叶片的相对含水量,降低叶片质膜透性和盐害对细胞膜的伤害,提高幼苗体内超氧化物歧化酶、过氧化物酶等细胞保护酶的活性,抑制过氧化作用产物丙二醛的积累;同时发现外源水杨酸和阿斯匹林还能够提高幼苗体内ＡＴＰ的含量,维持幼苗能量代谢和供应的正常进行,从而提高小麦对盐分胁迫的适应性     

ERF类转录因子OPBP1基因的超表达提高烟草的耐盐能力

ERF是植物中的一类重要的转录因子,参与调节植物的生长,发育以及抗胁迫等过程,对一烟草OPBP1基因（属于ERF类基因）的烟草转化,获得了该基因超表达的植株,转基因植株明显地增加了耐盐能力,Northern杂交结果表明,OPBP1基因有不同程度的表达,而且表达丰度与其耐盐性有一定的正相关性,凝胶阻滞实验结果证明OPBP1融合蛋白能特异地与含GCC盒的DNA序列结合,这些结果说明OPBP1基因可能作为一转录因子来调节烟草耐盐相关的基因。     

小麦盐胁迫应答cDNA的分离克隆及其特异片段SR07转烟草抗逆性分析

以宝丰7228小麦幼苗叶片为材料,利用DDRT-PCR技术对正常与盐胁迫条件下小麦叶片基因表达的差异进行分析后检测到27条差异cDNA片段,mRNA点杂交分析结果显示SR07片段存在明显的胁迫诱导表达特征。对SR07进行了序列测定和同源性对比分析后发现,SR07片段与植物水孔蛋白中质膜内在蛋白（PIPs）有87%的序列同源性。将SR07克隆到植物转化载体pCAMBIA中CaMV35S启动子下游,构建表达载体pCAMBIA-SR07,利用根癌农杆菌（Agrobacterium tumefaciens）质粒介导的遗传转化系统转化烟草（Nicotiana tabacum）,经筛选后获得3个烟草转化系。转化系的NaCl、PEG和甘露醇抗性实验结果表明,SR07转化株的抗盐性与对照株相比没有显著性差异（P〉0.05）,而PEG和甘露醇抗性与对照株相比有显著性差异（P〈0.05）,推测SR07表达蛋白可能在植物水分调节方面有重要作用。     

小麦盐胁迫相关基因的克隆与表达分析

采用RT-PCR方法,从小麦中克隆获得1个盐诱导小麦MYB类转录因子基因TaSIM（Triticum aestivum salt-induced MYB）,该基因cDNA全长1 213bp,具有1个831bp的开放阅读框,编码276个氨基酸,预测分子量约为29.903kD,等电点为10.12,推测的氨基酸序列中含有2个高度保守的SANT结构域。系统发生树分析表明,TaSIM与二穗短柄草XP₀03576185亲缘关系最近。半定量RT-PCR检测结果显示,TaSIM基因受盐胁迫诱导表达。亚细胞定位结果显示,TaSIM-hGFP融合蛋白定位于细胞核中。研究结果表明,小麦TaSIM基因编码的蛋白可能在细胞核内参与小麦对盐胁迫的应答反应。     

一氧化氮调节盐胁迫下小麦幼苗根部质膜H+-ATPase和焦磷酸酶活性提高耐盐性

采用外源一氧化氮(NO)供体硝普钠(SNP)研究了NO对盐胁迫下小麦(Triticum aestivum L.)幼苗耐盐性的影响.结果表明,0.1 mmol/L SNP处理显著缓解了1 50 mmol/L NaCl胁迫对小麦幼苗生长的抑制效应,包括水分丧失以及叶绿素降解,从而提高了小麦幼苗的耐盐性.进一步结合1 mg/mL血红蛋白处理则显著逆转了SNP诱导的上述效应;利用亚硝酸钠和铁氰化钾作为对照也证实了NO对小麦幼苗耐盐性的专一性调节作用,并可能与NO对小麦幼苗根部质膜H -ATPase和焦磷酸酶活性诱导有关.此外,尽管NO显著提高了盐胁迫下小麦幼苗根部细胞质膜H -ATPase和焦磷酸酶的ATP水解活性,但是对跨膜H 转运则没有明显影响.应用外源CaSO4和EGTA处理也证实,Ca2 可能在NO诱导的质膜H -ATPase和焦磷酸酶活性的提高过程中起信号作用.另外,分析盐胁迫下小麦幼苗根部Na 和K 含量的变化也发现,NO对Na 含量没有明显影响,但是却显著提高了K 水平和K /Na 比,这可能也是NO提高小麦幼苗耐盐性的原因之一.     

硫腺苷甲硫氨酸合成酶基因的克隆及其在盐胁迫条件下的不同表达

硫腺苷甲硫氨酸作为甲基供体在转甲基反应中起到重要作用.为了解硫腺苷甲硫氨酸在盐地碱蓬(Suaedasalsa (L.)Pall)耐盐中的作用,我们对可能编码硫腺苷甲硫氨酸合成酶的基因(SsSAMS2)进行了分析.该基因在经400 mmol/L NaCl处理的盐地碱蓬地上部分的λ-Zap cDNA文库中克隆到,其插入片段全长1 531 bp,包含一个395个氨基酸的开放阅读框架,该基因推断的分子量约为43 kD.SsSAMS2与长春花(Catharanthus roseus)的SAMS2在氨基酸水平上的一致性为93%.Southern杂交显示,SsSAMS2在盐地碱蓬基因组中可能是两个拷贝.Northern分析显示硫腺苷甲硫氨酸合成酶基因受NaCl等胁迫的正调控.酶活性检测表明,NaCl胁迫条件下该酶活性增强.