Proteomic comparison of four maize inbred lines with different levels of resistance to Curvularia lunata(Wakker)Boed infection

Protein profiles of leaves in four maize inbred lines with different disease resistance to pathogen Curvularia lunata(Wakker)Boed were studied by two-dimensional electrophoresis(2-DE)and mass spectrometry.Proteins were extracted from the forth leaf of maize seedlings 24 h after fungal inoculation,and fractionated by polyethylene glycol to precipitate the most abundant leaf protein,Rubisco,before gel separation.Protein profiles from 2-DE showed that total numbers of protein spots were increased in all four inbred lines inoculated with C.lunata CX-3 strain compared with the control.The numbers of changed protein spots in abundance were higher in resistant inbred lines than in susceptible ones,which implied that resistant inbred lines were more sensitive than susceptible ones to pathogen infection.Among proteins identified by MALDI-TOF MS,germin-like protein GLP and translation initiation factor eIF-5A were supposed to play important roles in maize resistance against C.lunata infection.     

Proteomic comparison of four maize inbred lines with different levels of resistance to Curvularia lunata （Wakker） Boed infection

Protein profiles of leaves in four maize inbred lines with different disease resistance to pathogen Curvularia lunata(Wakker)Boed were studied by two-dimensional electrophoresis(2-DE)and mass spectrometry.Proteins were extracted from the forth leaf of maize seedlings 24 h after fungal inoculation,and fractionated by polyethylene glycol to precipitate the most abundant leaf protein,Rubisco,before gel separation.Protein profiles from 2-DE showed that total numbers of protein spots were increased in all four inbred lines inoculated with C.lunata CX-3 strain compared with the control.The numbers of changed protein spots in abundance were higher in resistant inbred lines than in susceptible ones,which implied that resistant inbred lines were more sensitive than susceptible ones to pathogen infection.Among proteins identified by MALDI-TOF MS,germin-like protein GLP and translation initiation factor eIF-5A were supposed to play important roles in maize resistance against C.lunata infection.     

低温胁迫下克恩氏冬青幼苗叶片差异蛋白质分析

【目的】研究克恩氏冬青叶片应答低温胁迫的蛋白表达情况,探讨克恩氏冬青耐寒的作用机理。【方法】以2年生幼苗为试材,对其进行-8 ℃和-16 ℃低温胁迫处理,运用双向电泳技术(2-DE)结合质谱技术(MALDI-TOF/TOF MS)检测并分析叶片差异表达蛋白,并对差异蛋白进行了生物信息学分析。【结果】在低温胁迫下,克恩氏冬青幼苗叶片中蛋白表达发生了明显变化,胁迫前后发现共有31个显著差异的蛋白质点,经质谱检测及数据库检索,成功鉴定了其中23个差异表达蛋白点。对其中的20个蛋白成功进行了功能分类,主要涉及蛋白质和氨基酸代谢、光合作用、氧化还原平衡、防御作用、碳水化合物代谢、脂类代谢、氮素代谢等生物学过程。【结论】克恩氏冬青应答低温胁迫是多种蛋白质共同作用的结果,主要通过调节多种代谢过程中的相关蛋白表达来发挥作用。     

短日照处理诱导油松容器苗针叶差异表达蛋白质的功能分析

【目的】基于蛋白质水平探讨短日照处理调控油松(Pinus tabulaeformis)容器苗基础代谢和抗逆性机理的研究,完善油松容器苗的育苗和造林技术,为短日照处理育苗技术在我国北方地区困难立地造林中推广应用提供理论依据。【方法】以经日照长度为10 h、持续3 周短日照处理的油松容器苗针叶为研究对象,采用改良的酚法提取针叶蛋白。应用双向电泳结合二级质谱分析的蛋白质组学技术,研究短日照处理诱导油松容器苗针叶蛋白质表达的变化,分析差异表达蛋白质的鉴定和功能。【结果】成功获取5个短日照处理诱导表达差异显著的蛋白质,上调表达的是叶绿体放氧增强蛋白1(oxygen-evolving enhancer protein 1, OEE1)(蛋白点404)、甘油醛-3-磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase, GAPDH)(蛋白点539)、延伸因子-Tu(elongation factor-Tu, EF-Tu)(蛋白点654)、核酮糖-1,5-二磷酸羧化酶/加氧酶活化酶(ribulose bisphosphate carboxylase/oxygenase activase, RCA)(蛋白点681); 下调表达的是磷酸甘油酸激酶1(phosphoglycerate kinase 1, PGK1)(蛋白点641)。这些蛋白质的功能主要涉及光合代谢、糖代谢和胁迫防御。【结论】短日照处理诱导油松容器苗蛋白质差异表达,对油松容器苗的基础代谢和抗逆性产生重要影响。     

基于双向电泳技术的丙酮丁醇梭菌ATCC824不同产物阶段膜蛋白质组比较

建立丙酮丁醇梭菌在产酸阶段和产有机溶剂阶段的膜蛋白质组;并进行比较分析。研究采用差速离心法提取了该菌在产酸阶段和产有机溶剂阶段的膜蛋白,进行双向电泳分离后,对差异蛋白进行质谱分析。不同生长阶段膜蛋白质组存在明显差异,经质谱鉴定发现9个差异蛋白质,其中3个在产酸阶段高表达,6个在产有机溶剂阶段高表达。生物信息学分析表明,大部分蛋白含有明确的膜定位信息。产酸阶段高表达蛋白多与细胞信号转导有关,而产有机溶剂阶段高表达蛋白功能分布相对复杂。丙酮丁醇梭菌在不同产物阶段的差异膜蛋白被成功鉴定,它们可能在不同产物阶段的转换和细胞信号转导等过程中起到重要调控作用。     

玉米1,3,4-三磷酸肌醇5/6 激酶ITPK家族基因的鉴定和分析

 1,3,4-三磷酸肌醇5/6-激酶(ITPK)是一种在动物、植物、线虫中都比较保守的多功能酶, 在生物信号传导及生长发育中起重要作用。为了充分研究玉米中ITPK 家族基因系统分析、全生育期基因表达模式及逆境胁迫表达模式, 利用玉米基因组数据库, 通过生物信息学手段, 鉴定玉米ITPK 家族基因的全序列、定位和编码蛋白, 通过序列比对进行进化和分类分析。利用玉米高通量芯片表达数据进行组织表达差异性、干旱胁迫和病害胁迫表达谱分析。结果表明, 玉米基因组中含有6 个ITPK 家族基因, 分布于玉米的4 条染色体上。MEME 保守结构域分析显示, ZmITPK1-5 均含有3 个保守的ATP-grasp-4 结构域, ZmITPK6 含有两个ATP-grasp-4 保守结构域。进化树分析表明ZmITPK 可分为3 个亚家族。各个发育阶段中, 多数成员在生殖器官或营养器官中均有较高的表达量, 只有ZmITPK1 在所有器官中表达量都不高。ZmITPK2 和ZmITPK3 基因受干旱胁迫处理诱导不同程度上调表达。而在生物胁迫条件下均无显著上调或下调表达。     

HAK基因对玉米的遗传转化及其耐盐性研究

摘要：用农杆菌介导法将高亲和性钾离子转运体基因（HAK）和Bar基因转入5个优良玉米自交系7922、P138、265、238和271中,并对影响其遗传转化效率因素进行了优化．经PCR和RT-PCR检测证实获得阳性植株．除草剂涂抹实验证明Bar基因已经整合进玉米基因组．用不同浓度的盐溶液处理转基因植株和对照植株,发现转基因植株叶片中的K＋含量、脯氨酸和叶绿素含量均高于未转化植株,而Na＋含量低于未转化植株,表明HAK基因已经整合进玉米基因组,并通过过量表达提高了植株的耐盐性．     

橡胶树胶乳橡胶粒子死皮相关蛋白的鉴定及分析

橡胶树死皮是影响天然橡胶产量的重要因子之一。采用双向凝胶电泳对健康树和死皮树橡胶粒子中差异表达的蛋白进行分离,分析获得35个差异表达的蛋白点,通过基质辅助激光解析电离飞行时间质谱(MALDI-TOF MS)分析及搜索Uniprot rubber数据库后,有13个蛋白点被成功鉴定,其中橡胶延长因子(rubber elongation factor,REF)、法尼基焦磷酸合成酶(Ffarnesyl-diphosphate synthase,FPS)、谷胱甘肽过氧化酶(glutathione peroxidase,GPX)、谷胱甘肽还原酶(glutathione reductase,GR)、翻译控制肿瘤蛋白(translationally controlled tumor protein,TCTP)、热激蛋白(heat shock protein, HSP)等主要蛋白参与了橡胶的生物合成、活性氧代谢及细胞凋亡过程。说明橡胶的生物合成、活性氧代谢及细胞凋亡途径可能是橡胶树死皮发生的关键调控途径。     

水稻幼苗叶片应答稻瘟病侵染的差异蛋白谱分析

为了探讨水稻感病基因型种质响应稻瘟病侵染的蛋白质表达谱的变化规律和作用途径,以水稻感稻瘟病种质日本晴为材料,采用接种稻瘟病菌分生抱子悬浮液,24,48和72 h后提取叶片蛋白质,采用i TRAQ蛋白质组学技术研究稻瘟病胁迫下水稻叶片蛋白质组的变化.结果表明,稻瘟病侵染诱导了水稻幼苗叶片内涉及氧化还原平衡、防御、信号传导、糖和能量代谢、氨基酸代谢、光合作用,以及蛋白质代谢等代谢途径相关的53个蛋白质的表达量发生了改变.GO分析表明稻瘟病主要调控了植株体内细胞内平衡、代谢过程和蛋白质代谢等生物学过程.稻瘟病侵染激活了活性氧代谢、防御,以及热休克蛋白等相关的途径,而抑制了蛋白质生物合成过程.结合这些差异表达蛋白的丰度变化结合它们可能的功能,描绘了水稻应答稻瘟病侵染的蛋白质代谢网络,有助于在蛋白质水平上了解其应答过程.     

Proteomic analysis of long-term salinity stress-responsive proteins in Thellungiella halophila leaves

Salinity is one of the most severe environmental factors that may impair crop productivity. A proteomic study based on two-dimensional gel electrophoresis is performed in order to analyze the long-term salinity stress response of Thellungiella halophila, an Arabidopsis-related halophyte. Four-week-old seedlings are exposed to long-term salinity treatment. The total crude proteins are extracted from leaf blades, separated by 2-DE, stained with Coomassie Brilliant Blue, and differentially displayed spots are identified by MALDI-TOF MS or QTOF MS/MS. Among 900 protein spots reproducibly detected on each gel, 30 spots exhibit significant change and some of them are identified. The identified proteins include not only some previously characterized stress-responsive proteins such as TIR-NBS-LRR class disease resistance protein, ferritin-1, and pathogenesis-related protein 5, but also some proteins related to energy pathway, metabolism, RNA processing and protein degradation, as well as proteins with unknown functions. The possible functions of these proteins in salinity tolerance of T. halophila are discussed and it is suggested that the long-term salinity tolerance of T. halophila is achieved, at least partly, by enhancing defense system, adjusting energy and metabolic pathway and maintaining RNA structure.     

低磷胁迫对耐低磷玉米自交系幼苗光合特性的影响

采用沙培的方法,研究了低磷对来自细胞工程的耐低磷玉米自交系SD=C、SD-D和齐319叶片光合作用特征的影响. 结果表明：低磷胁迫下，玉米幼苗叶片的无机磷含量、光合速率下降. 耐低磷自交系SD-C、SD-D的无机磷含量、净光合速率下降幅度小于齐319. SD-C和齐319叶片光合速率的降低是受非气孔因素的限制，而SD-D则是受气孔因素的限制. 低磷处理使玉米叶片PSⅡ的光能转换和利用效率降低，耐低磷玉米自交系SD-C、SD-D的光能转换效率和利用效率高于齐319，受低磷胁迫的影响小于齐319. 玉米幼苗叶片内无机磷含量的差异可能是自交系SD-C、SD-D和齐319之间光合作用和叶绿素荧光产生差异的原因.     

Proteomic analysis of long-term salinity stress-responsive proteins in <Emphasis Type="Italic">Thellungiella halophila</Emphasis> leaves

Salinity is one of the most severe environmental factors that may impair crop productivity. A proteomic study based on two-dimensional gel electrophoresis is performed in order to analyze the long-term salinity stress response of Thellungiella halophila, an Arabidopsis-related halophyte. Four-week-old seedlings are exposed to long-term salinity treatment. The total crude proteins are extracted from leaf blades, separated by 2-DE, stained with Coomassie Brilliant Blue, and differentially displayed spots are identified by MALDI-TOF MS or QTOF MS/MS. Among 900 protein spots reproducibly detected on each gel, 30 spots exhibit significant change and some of them are identified. The identified proteins include not only some previously characterized stress-responsive proteins such as TIR-NBS-LRR class disease resistance protein, ferritin-1, and pathogenesis-related protein 5, but also some proteins related to energy pathway, metabolism, RNA processing and protein degradation, as well as proteins with unknown functions. The possible functions of these proteins in salinity tolerance of T. halophila are discussed and it is suggested that the long-term salinity tolerance of T. halophila is achieved, at least partly, by enhancing defense system, adjusting energy and metabolic pathway and maintaining RNA structure.     

补阳还五汤与高血压病气虚血瘀证“方证相关”的蛋白质组学研究

揭示补阳还五汤与高血压病气虚血瘀证"方证相关"的现代生物学基础,制作高血压病气虚血瘀证细胞模型。用补阳还五汤含药血清干预细胞模型,利用双向凝胶电泳(2-DE)及基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS/MS)技术筛选及鉴定差异表达的蛋白点。并对其进行生物学分析。结果高血压病气虚血瘀证组与健康对照组比较,差异蛋白质点有30个。其中,有16个蛋白上调,14个蛋白质下调。补阳还五汤组与高血压病气虚血瘀证组比较,差异蛋白质点有14个。其中,有9个蛋白上调,5个蛋白质下调。MALDI-TOF-MS/MS鉴定出:高血压病气虚血瘀证组与健康对照组差异蛋白点共有8个蛋白被成功鉴定出来。表达上调的蛋白有肽基脯氨酰异构酶A、肽基脯氨酸顺反异构酶A样1亚型、真核翻译起始因子5A-1 B亚型、微管蛋白beta-2C、CRA_b亚型、3-羟酰辅酶A脱氢酶2型异构体2。表达下调的蛋白有丙酮酸激酶同工酶M1亚型、抑制蛋白-1、抑制蛋白-1 1亚型、补阳还五汤组与高血压病气虚血瘀证组差异蛋白点共有3个蛋白被成功鉴定出来。表达上调的蛋白有丙酮酸激酶CRA_c亚型、热休克蛋白27;表达下调的蛋白有膜联蛋白A1,CRA_b亚型。这些蛋白多与促进或抑制细胞凋亡有关。说明高血压病气虚血瘀证存在着细胞凋亡,补阳还五汤可以纠正高血压病气虚血瘀证引起的细胞凋亡。这些差异蛋白可以作为高血压病气虚血瘀证的标志蛋白或补阳还五汤的作用靶点,抑制细胞凋亡可能是补阳还五汤与高血压病气虚血瘀证"方证相关"的现代生物学基础之一。     

锯缘青蟹对病原菌感染的急性反应功能蛋白质组

为研究锯缘青蟹对常见重要病原菌的急性反应蛋白质组．将锯缘青蟹随机分为四组．分别注射生理盐水、副溶血弧菌、鳗弧菌和嗜水气单胞菌．继而提取肌肉蛋白进行双向电泳．通过比较各组肌肉蛋白的双向电泳图谱获得三个差异表达的蛋白．对这三个差异蛋白进行肽质量指纹图谱及其生物信息分析．鉴定为Calexeitin、无翅蛋白片断、速激肽相关肽．这些结果对研究锯缘青蟹的抗病蛋白及其机理具有重要意义．     

适合杨树叶片蛋白质组学样品制备方法的比较

【目的】为了提高植物叶片中低丰度蛋白质的检测效率,建立有效的杨树叶片蛋白质组学分析的蛋白质提取方法。【方法】利用三氯乙酸-丙酮结合PEG分级提取方法和SDS-酚结合PEG分级提取方法对杨树叶片蛋白质进行提取,并采用SDS-PAGE和双向凝胶电泳(2-DE)对提取结果进行评价。【结果】SDS-PAGE实验结果显示,三氯乙酸-丙酮结合PEG方法优于SDS-酚结合PEG分级方法。三氯乙酸-丙酮结合PEG分级和非结合PEG分级提取蛋白质的双向电泳结果表明,PEG分级能有效地去除高丰度蛋白质Rubisco,提高低丰度蛋白质的检测。三氯乙酸-丙酮结合PEG分级法检测的蛋白质比非结合PEG分级法多90个蛋白质点。【结论】以LTQ-Orbitrap XL分析三氯乙酸-丙酮结合PEG分级分离的一些蛋白质显示,三氯乙酸-丙酮结合PEG分级对于2-DE和MS鉴定低丰度蛋白质是一种有效的分离方法。     

对B.t毒素敏感与抗性的昆虫细胞的差异蛋白质肽质指纹分析

采用2-DE技术分析了粉纹夜蛾TnH5对B.t CrglAC毒素敏感和抗性2种细胞总蛋白质,建立了TnH5细胞双向电泳图谱,获得了一些差异蛋白质.对部分差异蛋白质点采用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF/MS)测定了其胶内酶解后的肽质指纹图谱(PMF),查询数据库,结果表明,部分差异斑点分别与胞质外周蛋白、脂多糖生物合成蛋白、一种脱氢酶和类免疫球蛋白等类似.     

贵州西山系列玉米杂交种SSR标记纯度鉴定的研究

以11个贵州西山系列玉米杂交种及相应的15个亲本自交系为材料,采用SSR（Simple Sequence Repeat）分子标记方法,从60对多态性高的引物中分别筛选了双亲互补型特异引物,并利用特异引物进行玉米杂交种纯度鉴定的检验．     

玉米淀粉合成相关基因ZmSSIIa 的序列变异分析

 玉米淀粉合成酶ZmSSIIa 在淀粉生物合成途径中主要作用是将支链淀粉的分支链由6~10 个延长至12~25 个, 对于玉米籽粒产量和品质具有重要的影响。本研究在86 个优异玉米自交系中对该基因进行基因序列定点捕获, 并获得该基因全长为4836 bp 的核苷酸序列。基因多态性分析发现, 该基因座位上有78 个SNP 和19 个Indel。玉米ZmSSIIa 基因的编码区具有24个SNP, 将该基因划分成9 种编码区单倍型, 并编码7 种ZmSSIIa 蛋白质。供试材料的群体中, ZmSSIIa 基因至少经历8 次重组, 可能对单倍型的分化和连锁不平衡具有重要作用。中性进化测试表明该基因在供试群体中没有明显的人工选择。