寄主大豆对根结瘤的控制

我国９个大豆（ＧｌｙｃｉｎｅｍａｘＬ．Ｍｅｒｒ．）品种感染根瘤菌ＵＳＤＡ１１０后,产生不同的结瘤数,低者在２０个以下．高者在６０个以上。赤豆、绿赤豆也可被感染结瘤,而豇豆、扁豆则不能。超结瘤大豆ｎｔｓ３８２作为接穗时能诱导我国大豆原结瘤数有４５个的开育１０号、原结瘤数有１２个的大黄分别发生高结瘤。ｎｔｓ３８２作为砧木时,则不能表现超结瘤．表明超结瘤因子能传给我国大豆,反之存在于我国大豆中的限制超结瘤的因子也能传给ｎｔｓ３８２。ｎｔｓ３８２于ＮＯ３－环境中仍表现超结瘤的特点也能导入开育１０号、大黄及赤豆根部,并使之在ＮＯ３－环境中结瘤。在ＮＯ３－环境中不能结瘤的开育１０号作为接穗,ｎｔｓ３８２作为砧木的嫁接植株,于子叶生长阶段接受ＮＯ３－时,仍能结瘤,于真对生长时接受ＮＯ３－时．则不能结瘤,表明限制结瘤因子于真叶细胞中被诱导形成。     

大豆突变基因的遗传分析及窄叶突变基因的ＲＡＰＤ标记

宽叶、无4粒荚大豆品种鲁豆4号（代号LD4）经烷化剂EMS诱变处理选育出窄叶和4粒荚同时发生突变的稳定突变系E182。突变系与亲本杂交遗传分析表明,“宽叶与窄叶”和“无4粒荚与有4粒荚”2对性状均符合孟德尔单因子的31分离比率,说明窄叶和4粒荚突变性状均受单个隐性核基因控制。F     

大豆种子萌发过程中的差异蛋白质组研究

运用蛋白质组学技术对大豆(Glycinemax)N2899种子萌发0h、8h、36h、60h4个时期蛋白质的差异表达情况进行了研究.结果发现,在考马斯亮蓝染色的双向电泳pH3～10胶上,PDQuest图像分析软件可识别的点约350个,其中表达量变化2.5倍以上的蛋白质点有24个,而绝大部分大豆种子贮藏蛋白在萌发期尚未降解.在萌发的第一阶段,24个差异表达蛋白中有10个蛋白质的丰度发生变化.第二阶段,差异表达蛋白的种类和量增加,其中15个蛋白质是动态变化的,14个蛋白质在胚根突破种皮时表达量达到峰值,表明吸胀后种子内的生命活动越来越强.对这24个蛋白质点进行胶内酶解,用基质辅助激光解析电离飞行时间质谱测定均获得肽质量指纹图谱.搜索大豆的UniGene库初步鉴定出6个蛋白质,分别是核苷二磷酸激酶、热激蛋白、硫氧还蛋白、35ku种子成熟蛋白及种子成熟蛋白PM36.对这些蛋白质在种子萌发过程中可能的作用进行了讨论.     

Quantitative Proteomic Analysis of Low Linolenic Acid Transgenic Soybean Reveals Perturbations of Fatty Acid Metabolic Pathways

To understand the effect of fatty acid desaturase gene (GmFAD3) silencing on perturbation of fatty acid (FA) metabolic pathways, the changes are compared in protein profiling in control and low linolenic acid transgenic soybeans using tandem mass tag based mass spectrometry. Protein profiling of the transgenic line unveiled changes in several key enzymes of FA metabolism. This includes enzymes of lower abundance; fabH, fabF, and thioestrase associated with FA initiation, elongation, and desaturation processes and LOX1_5, ACOX, ACAA1, MFP2 associated with β‐oxidation of α‐linolenic acids pathways. In addition, the GmFAD3 silencing results in a significant reduction in one of the major allergens, Gly m 4 (C6T3L5). These results are important for exploring how plants adjust in their biological processes when certain changes are induced in the genetic makeup. A complete understanding of these processes will aid researchers to alter genes for developing value‐added soybeans.     

脲酶抑制剂氢醌对大豆结瘤、固氮和木质部溶质氮形态的影响

首次报导了在白浆土和砂壤土上使用氢醌(HQ)抑制土壤脲酶活性,有效地缓解尿素分解产物及其随后的氧化产物对大豆结瘤和固氮活性的抑制效应,结果表明:1.HQ浓度在40PPM以内对大豆幼苗生长及初生结瘤表现促进作用;进一步提高HQ浓度将使大豆根系生长受阻变态而阻止结瘤。2.HQ(10—50PPM)提高了离体活性大豆根瘤类苗体悬液的耗氧量(79.4—86.1％)和琥珀酸脱氢酶活性(124.7—138.4％)。3.盆栽和田间试验证实,由于HQ缓解了尿素的分解,从而颇大减轻了尿素对大豆结瘤和固氮(乙炔还原活性)的抑制效应;通过大豆木质部中溶质氮形态(酰胺、酰脲和硝酸盐)的分析进一步证实了,大豆植株从根部向地上部运输的氮素形态同土壤氮转化强度和根瘤固氮强度(酰脲相对丰度)之间的紧密联系。4.由于麦秸还田土壤脲酶活性提高,故应提高HQ剂量;与此同时,通过麦秸的“氮因子效应”便能完全解除尿素对大豆结瘤固氮的抑制,并为大豆籽实发育提供了丰富的土壤氮源。     

Competitiveness of indigenous populations of Bradyrhizobium japonicum serocluster 123 as determined using a root-tip marking procedure in growth pouches

Soil Bradyrhizobium populations limit nodule occupancy of soybean by symbiotically-superior inoculant strains throughout much of the American midwest. In this study, the competitiveness of indigenous populations of B. japonicum serocluster 123 from Waukegan and Webster soils was evaluated in growth pouches using a root-tip marking procedure. The native rhizobia were from soils incubated 0–8 h in soybean root exudate (SRE) or plant nutrient solution (PNS) prior to inoculation. Populations of serocluster 123 strains in soil and nodule occupancy by these strains were assessed using fluorescent antibodies prepared against B. japonicum USDA 123. There were no significant differences in populations that came from SRE or PNS incubated soils: both populations increased in number over the incubation period. Nodule occupancy by both populations in growth pouches was similar to that previously encountered in field studies with these two soils. With the Waukegan soil, the serocluster 123 population dominated nodulation forming 69 and 62% of taproot nodules above and below the root tip mark, respectively. However, for the more alkaline Webster soil, serocluster 123 strains were much less competitive, producing only 9 and 13%, respectively, of the nodules formed above and below the root tip mark. In growth pouches, soil populations of bradyrhizobia from the Webster soil produced significantly more nodules than those from the Waukegan soil, but both strains and a pure culture of USDA 110 had a similar distribution of nodules.     

小麦耐盐突变体盐胁迫下的蛋白质组分析

首次采用双向电泳的方法分析1％NaCl胁迫72h的一对小麦耐盐(RH8706-49)及敏盐突变体(H8706-34)的蛋白质组。经过MALDI-TOF分析和数据库检索发现两者在H^ -ATP酶β亚基、谷氨酰胺合成酶前体、OEC33和RuBP羧化酶小亚基等5个候选蛋白存在质或量的差异。这5种蛋白均为叶绿体蛋白,它们很可能在盐胁迫下对维持叶绿体及整个细胞的功能起到重要作用。     

拟南芥耐盐突变体stg2的筛选

采取在高盐平板上萌发的方法,对一个雌激素诱导激活型拟南芥突变体库进行了耐盐突变体的筛选,最终得到了2株稳定的耐盐突变体。本文中对其中的一株耐盐突变体,命名为stg2(salt tolerance during germination 2),进行了研究。遗传实验表明它的耐盐特性是受雌激素诱导的,是功能获得型的耐盐突变体。本实验中还探讨了stg2突变体的筛选过程及耐盐生理特点。     

OsPT6基因过表达对低磷条件下菜用大豆结瘤及固氮的影响

采用砂培方法,以转OsPT6基因的菜用大豆(T3株系)与其非转基因(NT)受体品种为实验材料,研究了两者在低磷条件下的生长发育指标,植株有效磷、全磷、全氮、豆血红蛋白和籽粒蛋白质含量以及谷氨酰胺合成酶活性的差异,并对植株结瘤及固氮相关基因表达进行检测,为阐明转OsPT6基因菜用大豆在低磷条件下结瘤及固氮相关机理提供理论依据。结果显示:(1)转基因植株的株高、茎粗、花数和荚数、根瘤数均显著高于NT植株。(2)转基因植株根、茎、叶及根瘤中有效磷,全株总磷、总氮含量,根瘤中的豆血红蛋白含量、功能叶片中谷氨酰胺合成酶的活性和籽粒蛋白质含量均显著高于NT植株。(3)相关性分析表明,豆血红蛋白含量、谷氨酰胺合成酶活性、总磷、总氮含量4个指标间均呈显著正相关关系。(4)GmENOD40a、GmENOD40b、GmGS1β1、GmGS1β2基因在转基因植株中的表达量显著高于NT植株。研究表明,OsPT6基因过表达增强了菜用大豆在低磷条件下的结瘤及固氮能力,该研究结果为进一步研究其调控机理奠定了基础。     

水稻短根毛突变体Ossrh2的表型分析与基因定位

在粳稻品种中花11为遗传背景的T-DNA突变体库中筛选获得一个遗传稳定的水稻（Oryzasativa）短根毛突变体Ossrh2（Oryza sativa short root hair2）。突变体在苗期表现为根毛数量减少,为野生型的61.4%,根毛长度明显变短,只有野生型的22.8%,同时根毛增粗,根毛形态也发生了变异,局部扭曲膨胀和分叉,除此之外突变体的地上部和根部生长情况与野生型相比没有显著差异。遗传分析表明,该突变性状受1对隐性单基因控制。通过对突变体T2和F2代的分子检测发现,该突变体表型非T-DNA插入引起。利用Ossrh2纯合体和籼稻品种Kasalath杂交构建的F2群体对OsSRH2进行基因定位,发现其与第10号染色体短臂上的SSR（simple sequence repeat）标记RM6370和RM474连锁,遗传距离分别为1.1cM和3.0cM。通过在两标记间发展3个新的STS（sequence-taggedsite）标记,将OsSRH2基因定位于标记S1227和S1531之间,物理距离约为304kb,为进一步克隆OsSRH2打下了基础。     

A nopA Deletion Mutant of Sinorhizobium fredii USDA257, a Soybean Symbiont, is Impaired in Nodulation

Sinorhizobium fredii USDA257 employs type III secretion system (T3SS) to deliver effector proteins into the host cells through pili. The nopA protein is the major component of USDA257 pili. The promoter region of USDA257 nopA possesses a well conserved tts box. Serial deletion analysis revealed that the tts box is absolutely essential for flavonoid induction of nopA. Deletion of nopA drastically lowered the number of nodules formed by USDA257 on cowpea and soybean cultivar Peking. In contrast to the parental strain, the USDA257 nopA mutant was able to form few nodules on soybean cultivars McCall and Williams 82. Light and transmission electron microscopy examination of these nodules revealed numerous starch grains both in the infected and uninfected cells.     

Primary Analysis of Components in Different Soybean Nodulating Mutants and their Effects on Nitrate Reductase Activity and Nodulation

The water extracts of leaves and roots from supernodulating soybean (Glycine max (L.) Merr. ) nts 382 and nonnodulating soybean Nod 49 have been chromatographed using filtering method through the column (25 cm × 2 cm) Sephadex G25 and 4 fractions, namly, nts 382 (Nod 49) F1, nts 382 (Nod 49) F2, nts 382 (Nod 49) F3, and nts 382 (Nod 49) F4 could be distinguished according to nitrate reductase (NR) activities inhibited by the eluate. The inhibition of NR activity by the noninoculated nts 382 F2 and the nts 382 F4 in vitro were much stronger than that by the inoculated nts 382 F2 and nts 382 F4. On the contrary, the obvious inhibition of NR activity in vitro by the noninoculated Nod 49 F2 and Nod 49 F4 were substantialy strengthed again by the innoculated Nod 49 F2 and Nod 49 F4. The facts indicated that the quantity of NR inhibitors in the leaf cells of soybean nts 382 reduced after the inoculation but was that in the inoculated Nod 49 leaf cells further more accumulated. Both nodulations assays, the nodulation of soybean "Bragg " injected with inoculated nts 382 Fl, nts 382 F2, nts 382 F3 and nts 382 F4 from leaves and roots and the nodulation of soybean nts 382 injected with inoculated Nod 49 F2, Nod 49 F3 and Nod 49 F4 from leaves only showed that nts 382 Fl and nts 382 F2 increased nodules of soybean "Bragg" by 1 to 3 times but nts 382 F3 and nts 382 F4 did not. Inhibition of soybeannts 382 nodulation by inoculated Nod 49 F2 Nod 49 F3 and Nod 49 F4 expressed that the Nod 49 F4 only inhibited the nodulation strongly by one time in the experiments with nts 382 plants with leaves, and by 15 times in the experiments with nts 382 plants without leaves at 10 d of inoculation and injection and this inhibition was nonreversible even after stopping injection from the 11th day to the 15th day after inoculation.     

Comparative proteomic analysis of soybean nodulation using a supernodulation mutant, SS2-2

Legumes have the ability to form root nodules that fix atmospheric nitrogen through a symbiotic interaction with nitrogen-fixing bacteria. As a first step in dissecting the molecular process of nodulation, proteome reference maps of soybean roots and nodules were constructed. Time course analysis revealed that the transition from root to nodule was accompanied with downregulation of defense-response related proteins, including Mn-superoxide dismutase, peroxidase (Prx), PR10, and stress-induced protein, leading to the initiation of a symbiotic interaction between the two partners. Following nitrogenase biosynthesis, the host plant cooperated with the rhizobia to fix atmospheric nitrogen under microaerobic conditions via expression of leghemoglobins and antioxidant proteins. Comparative proteome analysis indicated lower expression of malate dehydrogenase (MDH), leghemoglobins and nitrogenase in the nodule development of the supernodulation mutant, SS2-2, as compared to the wild type, indicating that SS2-2 forms functionally immature nodules in higher numbers with the lower activity of nitrogen fixation.     

水稻无侧根突变体的根向重力性异常

用化学诱变剂 (NaN3 )处理粳稻品种大力 (O ryzasativaL .cv .Oochikara) ,得到具有 2 ,4 D抗性、无侧根和根向重力性异常的突变体RM 10 9。对原品种为父本和突变体为母本的杂交后代F1、F2 根向重力性的遗传分离进行了研究。结果表明 :突变体的根向重力性异常 ,其性状是单显性基因控制且不受光照和黑暗培养的影响。通过对根冠组织切片观察发现 :突变体根冠中含淀粉体的细胞数量比大力少 ,根冠细胞中淀粉体的直径为原品种的 5 0 %且集中排列于细胞内的一角 ,其排列沉积方向与重力方向相同。推测 :突变体的根向重力性异常与淀粉体直径变小有关     

蛋白质组双向电泳实验中一些常见失误的分析

从以固相pH梯度等电聚焦为第一向,SDS-PAGE电泳为第二向进行的蛋白质组双向电泳实验中挑选了一些常见的硝酸银染色2-DE失误图谱,将它们进行分类,初步分析造成各类失误的可能原因,探讨相应的解决方法和总结实验操作中应该注意的事项．     

野生大豆与栽培大豆种子差异蛋白质组学研究

运用蛋白质组学方法比较研究3个野生大豆(Glycinesoja)和3个栽培大豆(Glycinemax)的种子贮藏蛋白差异情况.结果发现,在考马斯亮蓝染色的双向电泳pH4～7的胶上,经过PDQuest图像分析软件平均可检测到550个左右的蛋白质点.进一步分析发现,表达量变化2.5倍以上的点有10个,其中大部分蛋白质仅在栽培大豆中检测到.对这10个蛋白质点进行了胶内酶解,用基质辅助激光解析电离飞行时间质谱测定均得到了肽质量指纹图谱.搜索大豆UniGene库和NCBI库共鉴定出5个蛋白质,主要是与大豆抗性、抗营养以及种子萌发相关的蛋白质,包括大豆血凝素,种子成熟蛋白PM24,糖结合蛋白,胰蛋白酶抑制剂p20以及成熟多肽.对这些蛋白质可能的作用进行了讨论.