苜蓿属3种不同花色基因型的染色体核型分析

基于形态学的显著差异,对紫花苜蓿(Medicago sativa L.)、黄花苜蓿(Medicago falcata L.)和白花苜蓿(Alfalfa with Cream flower)进行染色体核型分析,结果表明,3种类型的苜蓿材料在染色体核型方面有显著差别,白花苜蓿和黄花苜蓿都是随体染色体,其中,黄花苜蓿有2条为端部着丝点染色体,其核型公式分别为:紫花苜蓿2n=4x=24m+8sm、黄花苜蓿2n=4x=24m+6sm+2T(SAT)、白花苜蓿2n=4x=24m+8sm(SAT)。紫花苜蓿与白花苜蓿的染色体具有89%的相似性,只是在紫花苜蓿7号染色体和白花苜蓿15号染色体存在随体有无的区别;黄花苜蓿染色体具有独特性,显著区别于紫花苜蓿和白花苜蓿,但从其19号染色体来看,参与了紫花苜蓿或白花苜蓿的构建;白花苜蓿15号随体染色体与黄花苜蓿25号随体染色体和紫花苜蓿7号染色体有高度的相似性,但也存在随体有无的区别。     

牛角花齿蓟马为害对苜蓿无性系根茎叶及同化产物分配的影响

为探索同化产物分配利用与苜蓿耐蓟马的关系,本试验以扦插的抗蓟马苜蓿无性系R-1和感蓟马苜蓿无性系 I-1为材料,研究不同虫口牛角花齿蓟马为害对苜蓿的抗性、根、茎和叶生长特性及可溶性糖含量的影响.结果表明: 随着虫口压力的增大,R-1和I-1苜蓿的受害指数升高;在相同虫口压力下,R-1苜蓿的受害指数显著低于I-1.受蓟马为害后,R-1和I-1苜蓿株高降低、叶面积减少、茎秆变细、节间长变短、节间数增加,根颈和主根直径加粗、侧根增多.在低虫口密度下,随虫口压力增大,R-1和I-1苜蓿地上部生物量增加,根冠比下降,分配到茎的生物量比例升高;在高虫口密度下,地上部生物量随虫口压力增大而减少,根冠比增加,分配到根系的生物量比例升高;R-1根冠比和茎生物量比例随虫口压力变化曲线的拐点均为每枝条5头,I-1根冠比和茎生物量比例随虫口压力变化曲线的拐点均为每枝条3头.在低虫口压力下,R-1苜蓿茎和叶中的可溶性糖含量随虫口压力增加而升高;在高虫口压力下,茎和叶中的可溶性糖含量随虫口压力增加而下降;根中可溶性糖含量随虫口压力增加持续下降.I-1根、茎和叶中的可溶性糖含量均随虫口压力增加持续下降.牛角花齿蓟马为害后,R-1根、茎和叶的农艺性状及抗性比I-1好,对同化产物的分配利用率高.     

龙牙楤木总皂苷含量测定及其抗炎镇痛活性评价

建立了一种龙牙楤木中总皂苷的含量测定方法,比较了龙牙楤木根皮和茎皮中总皂苷含量,并研究了龙牙愡木的抗炎镇痛作用。采用超声辅助提取,用紫外-可见分光光度法,以齐墩果酸为对照品建立标准曲线;以5%香草醛-冰醋酸为显色剂,检测波长550nm,用比色法测定总皂苷含量。结果显示所建立的方法稳定可靠,龙牙楤木不同部位皂苷含量差异较大,根皮总皂苷含量为43.50mg/g,而茎皮为24.38mg/g。总皂苷在0.02~0.10mg/mL(R~2=0.999 6)质量浓度范围内呈良好线性关系,根皮和茎皮的平均回收率分别为100.12%(RSD=1.10%)和99.84%(RSD=1.77%)。采用醋酸致小鼠扭体试验和耳廓肿胀试验研究龙牙楤木总皂苷抗炎镇痛活性,结果表明龙牙楤木总皂苷能显著减轻醋酸对试验小鼠内脏所致的疼痛(P<0.05)和降低由二甲苯所致实验小鼠耳廓的肿胀程度(P<0.05),显示出较强的抗炎镇痛作用,且根皮的作用效果比茎皮好。龙牙愡木抗炎镇痛活性可能与其总皂苷含量呈正相关。建立的总皂苷含量测定方法简便、准确、重现性好,可作为龙牙楤木根皮与茎皮中总皂苷的含量测定方法。     

苜蓿根际土壤水提液化感作用的研究

以10个国内外引进的紫花苜蓿品种为材料,研究其不同浓度根际土壤水提液对受体苜蓿阿尔冈金种子发芽、幼苗生长和幼苗鲜重的影响.结果表明：不同品种紫花苜蓿根际土壤水提液对受体苜蓿种子发芽都存在抑制作用,且随着水提液浓度增大抑制作用有增强的趋势,并以爱维兰、牧歌401＋Z和牧野水提液的抑制作用较显著;大部分苜蓿品种的根际土壤水提液对受体苜蓿幼苗的生长存在抑制作用,但对根的生长具有促进作用,对幼苗茎叶和根生物量的积累均具有促进作用,且对茎叶的促进作用大于根,并以全能＋Z、射手、牧歌401＋Z和爱维兰的作用较明显.研究表明,苜蓿根际土壤水提液的化感作用在品种间和浓度间存在显著差异,受体苜蓿各器官对化感作用的反应也不同.     

鳢肠不同部位抗骨质疏松活性及化学成分比较研究

为了比较鳢肠不同部位提取物的抗骨质疏松活性及总皂苷、总黄酮含量,采用MTT法和对硝基苯酚磷酸二钠法测定鳢肠不同部位乙醇提取物对UMR106细胞的增殖和碱性磷酸酶(ALP)活性;采用紫外分光光度法测定鳢肠不同部位总皂苷和总黄酮含量。结果表明,鳢肠根和叶提取物对UMR106细胞的增殖具有促进作用,浓度为50μg/mL时促增殖作用最强,增值率分别为7.7%和16.9%,茎提取物没有明显的作用;鳢肠根、茎、叶提取物对ALP活性均有促进作用,浓度为100μg/mL时促增殖作用最强,分别促进18.9%、20.1%和17.0%;鳢肠根、茎、叶中总皂苷含量分别为3.6%、4.0%和2.2%,总黄酮含量分别为1.4%、1.4%和3.4%。这些结果表明;鳢肠不同部位提取物均有抗骨质疏松活性,其不同部位总皂苷和总黄酮的含量差异较大。     

不同施钾水平对苜蓿营养物质及抗蓟马性的影响

【目的】明确施钾是否能有效提高苜蓿对蓟马的抗性以及钾-营养物质-抗虫性之间的关系。【方法】以感蓟马紫花苜蓿Medicago sativa品种甘农3号(Gannong No.3)和抗蓟马紫花苜蓿品种甘农9号(Gannong No.9)为材料,设0,6,9,12和15 g/m2(K2O)5个钾水平,在大田蓟马为害高峰期,评价和测定了不同钾水平处理下苜蓿的受害指数、虫口数量、叶片钾含量、碳氮比和游离氨基酸变化。【结果】在第2茬中,随着钾水平的升高,甘农3号和甘农9号老叶和心叶的钾含量上升,可溶性糖和碳氮比升高,游离氨基酸含量下降,虫口数量无显著性差异,受害指数显著降低(P0.05),且在9 g/m2(K2O)水平下最低。在第3茬中,甘农3号和甘农9号的受害指数、叶片钾含量、碳氮比及游离氨基酸的变化规律和第2茬相同,但虫口数量上升。第2茬和第3茬中,不同施钾水平下,苜蓿叶片钾含量与受害指数负相关但不显著(P0.05)。施钾后甘农3号的受害指数均低于未施钾甘农9号的受害指数。【结论】钾元素可通过提高苜蓿碳氮比,降低游离氨基酸的含量来提高苜蓿对蓟马的耐害性。在大田条件下,通过施钾管理来提高苜蓿品种对蓟马的耐害性是一种有效的措施。     

HPLC-DAD和UPLC-MS/MS对蔓花生中二苯乙烯类化合物的研究

采用HPLC-DAD和UPLC-MS/MS对蔓花生中白藜芦醇、白藜芦醇苷和二苯乙烯苷三种二苯乙烯类化合物进行鉴定,建立HPLC-DAD同时测定三种化合物含量的方法。HPLC-DAD分析采用依利特ODS1色谱柱,柱温40℃,流动相A为甲醇,B为1%乙酸水溶液,流速1.0 mL/min。UPLC-MS/MS采用ACQUITY UPLC BEH C18色谱柱,流动相A为乙腈,B为水,电喷雾离子源,负离子检测,数据采集方式为多反应监测模式(MRM)。结果表明:蔓花生根、茎和叶中白藜芦醇含量分别为66.28、90.83和81.56μg/g,白藜芦醇苷含量分别为123.78,302.77和236.53μg/g,根和茎中二苯乙烯苷含量分别为673.60和764.65μg/g,叶中未检测到二苯乙烯苷。     

苜蓿种子萌发和幼苗生长对温度、光照和埋深的响应

紫花苜蓿(Medicago sativa)是世界人工草地重要的优良牧草。研究其种子萌发和幼苗生长对温度、光照及埋深等环境因子的响应,对苜蓿草地的种植及管理具有重要的实践指导意义。采用室内控制实验,分析了紫花苜蓿种子萌发和幼苗生长对恒温(10、15、20、25、30℃)、变温(6/15、10/20、15/25、15/30、20/30℃)、光照(12 h光照/12 h黑暗、24 h黑暗)以及埋深(1、2、3、4、5 cm)等环境因子的响应特征。结果表明,苜蓿种子的萌发温度范围宽,在供试的10种恒温和变温下萌发率为50%~88%,且温度处理对苜蓿种子的萌发率没有显著的影响;黑暗对苜蓿种子萌发有抑制作用,12 h光照/12 h黑暗条件下,苜蓿种子萌发率为69%~88%,而24 h全暗条件下为50%~77%,其中10、15、30、10/20和15/30℃温度下,光照处理均显著高于全暗处理。埋深1~5 cm的苜蓿种子出苗率没有显著差异,且幼苗株高、干重、鲜重均没有受到显著影响;埋深对苜蓿根生长存在显著影响,埋深1~2 cm根长度及重量均显著高于其他埋深处理。以上结果表明,苜蓿种子萌发的温度范围宽,恒温和变温下都有较高的萌发率,适合播种季节要求宽泛;黑暗显著抑制紫花苜蓿种子的萌发,田间种植苜蓿的最佳埋深为1~2 cm。     

天蓝苜蓿单细胞培养植株再生

采用了改良K8p和Ay3两种培养基,对天蓝苜蓿进行单细胞培养并得到再生植株。来自天蓝苜蓿胚轴愈伤组织的细胞系,在单细胞培养中的愈伤组织分化率明显高于来自子叶愈伤组织的细胞系。改良K8p培养基较之Ay3培养基更利于天蓝苜蓿单细胞培养。生物素、泛酸钙和葡萄糖对细胞系的细胞分裂、愈伤组织诱导和分化有促进作用。赤霉素促进再生幼芽向植株发育。     

响应面法优化超声提取苜蓿皂苷工艺条件

目的:利用响应面法对超声提取苜蓿皂苷的工艺条件进行了优化.方法:研究了超声波条件下影响提取的几个因素,包括超声时间、超声温度、超声功率、固液比等,并通过响应面法优化工艺条件.结论:根据中心组合设计原理采用四因子三水平的响应面分析法,通过对各因子显著性和交互作用的分析,得出了超声提取苜蓿皂苷的最佳工艺条件为:超声时间22min,超声温度43℃,超声功率403W,固液比1:51(g/ml),此时苜蓿皂苷的得率为4.53%.     

Population genetic structure of Sinorhizobium meliloti and S. medicae isolated from nodules of Medicago spp. in Mexico

We studied the genetic structure of 176 bacterial isolates from nodules of Medicago sativa, M. lupulina and M. polymorpha in fifteen sites distributed in three localities in Mexico. The strains were characterized by multilocus enzyme electrophoresis, plasmid profiles, PCR restriction fragment length polymorphism of 16S rRNA genes and of the intergenic spacer between 16S and 23S rRNA genes, and partial sequences of glnII, recA and nodB. Most of the strains were classified as Sinorhizobium meliloti, and a high genetic diversity was recorded. Six strains were classified as Sinorhizobium medicae, with no genetic variation. Phylogenetic and population genetic analyses revealed evidence of frequent recombination and migration within species.     

Phylogeny and character evolution in Medicago (Leguminosae): Evidence from analyses of plastid trnK/matK and nuclear GA3ox1 sequences

? Premise of the study: The genus Medicago, with about 87 species, includes the model legume species M. truncatula, and a number of important forage species such as M. sativa (alfalfa), M. scutellata (snail medic), and M. lupulina (black medic). Relationships within the genus are not yet sufficiently resolved, contributing to difficulty in understanding the evolution of a number of distinguishing characteristics such as aneuploidy and polyploidy, life history, structure of cotyledons, and number of seeds per fruit. ? Methods: Phylogenetic relationships of 70-73 species of Medicago and its sister genus Trigonella (including Melilotus) were reconstructed from nucleotide sequences of the plastid trnK/matK region and the nuclear-encoded GA3ox1 gene (gibberellin 3-β-hydroxylase) using maximum parsimony and Bayesian inference methods. ? Key results: Our results support certain currently recognized taxonomic groups, e.g., sect. Medicago (with M. sativa) and sect. Buceras. However, other strongly supported clades-the "reduced subsection Leptospireae clade" that includes M. lupulina, the "polymorpha clade" that includes M. murex and M. polymorpha and the "subsection Pachyspireae clade" that includes M. truncatula-each of which includes species presently in different subsections of sect. Spirocarpos, contradict the current classification. ? Conclusions: These results support the hypothesis that some characters considered important in existing taxonomies, for example, single-seeded fruits that have arisen more than once in both Medicago and Trigonella, are indeed homoplastic. Others, such as the 2n = 14 chromosome number, have also arisen independently within the genus. In addition, we demonstrate support for the utility of GA3ox1 sequences for phylogenetic analysis among and within closely related genera of legumes.     

冷蒿和苜蓿对意大利蝗生长及生殖力的影响

以紫花苜蓿 Medicago sativa (L.)、冷蒿 Artemisia frigida Willd.Sp.Pl.以及二者1:1混合饲喂意大利蝗Calliptamus italicus (L.)一龄蝗蝻,冷蒿饲喂不能完成其生活史,两种食料混合饲喂的发育进度要快于紫花苜蓿饲喂的个体。以三种食料分别饲喂意大利蝗成虫,研究其生长及生殖力的变化,结果表明：与取食紫花苜蓿和取食混合食料相比,取食冷蒿抑制了意大利蝗成虫体长、体重、雌雄成虫寿命、交配率和单雌产卵量,导致交配期提前,雌虫死亡率升高,而对卵囊内卵粒数无影响。与取食紫花苜蓿相比,取食混合食料促进了卵囊内卵粒数的增加,其余指标没有显著变化。这些结果表明两种牧草的隔离种植对减轻意大利蝗为害是十分必要的。     

紫花苜蓿化学成分的研究(英文)

为研究紫花苜蓿（Medicago scttiva L．）地上部位的化学成分,通过溶剂萃取、硅胶柱色谱、凝胶柱色谱分离纯化,从紫花苜蓿地上部分分离得到9个化合物,根据质谱和核磁共振数据鉴定为小檗碱（1）、大黄素（2）、大黄素8-O-β-D-葡萄糖苷（3）、soyαsαpogenol B-3-O-β-glcA（4）、邻羟基苯甲酸（5）、反式对羟基肉挂酸（6）、顺式对羟基肉桂酸（7）、正三十烷醇（8）和β-胡萝卜苷（9）。其中化合物1—4为首次从苜蓿属植物中分离得到。     

Comparison of nucleotide diversity and symbiotic properties of Rhizobium meliloti populations from annual Medicago species

Abstract: Forty-three isolates of Rhizobium meliloti were trapped from soil with five annual species of Medicago (M. polymorpha, M. truncatula, M. rigidula, M. orbicularis and M. minima ) and one perennial species of Medicago (M. sativa) . The annual species were growing naturally near the soil sampling site, and the commonly studied perennial species was used for comparison. Each R. meliloti was characterized by PCR-RFLP methods applied to two DNA regions nested between 16S rRNA and 23S rRNA genes and between nif D and nif K genes. They fell into two highly divergent groups (groups I and II), separated at a genetic distance of 0.024 by rDNA-amplified pattern analysis (profiles R1 and R2) and at 0.029 by nif -amplified pattern analysis (profiles N1-N2 and N3). These two groups were consistent with some cross-nodulation and -fixation results: rhizobia with the R1 genetic background elicited rudimentary nodules and could not fix nitrogen on M. polymorpha , while they were able to nodulate the five other species of Medicago . In contrast, rhizobia with an R2 profile were highly effective on M. polymorpha and poorly nodulated M. rigidula species, but were able to nodulate efficiently the other species. The striking phenotypic traits on M. polymorpha were also shared by reference strains: strains genetically closed to R2 type triggered typical and efficient nodules on M. polymorpha while those close to R1 type elicited rudimentary and non-efficient ones. Our results suggest that the presence of R. meliloti with R2 genetic backgrounds could be favoured by the distribution of M. polymorpha species.     

Genome size and base composition in Medicago sativa and M. truncatula species.

The genome size (1C value) and base composition of 14 ecotypes of two species of tetraploid and diploid Medicago have been assessed by flow cytometry. These parameters vary both between and within species. The diploid annual Medicago truncatula Gaertn. had the smallest genome of the group studied (which also covered M. sativa L. subsp. sativa, M. sativa L. subsp. caerulea (Less. ex Ledeb.) Schmalh., M. sativa L. subsp. quasifalcata Sinsk., M. sativa L. subsp. x varia (Martyn) Arcangeli; however, its ecotypes revealed substantial intraspecific variation. The smallest M. truncatula genome observed was ecotype 108-1 with 1C = 0.49 pg and 38.1% GC and the largest was Jemalong with 1C = 0.57 pg and 38.6% GC. The degree of polysomaty in these Medicago was low, although in some tissues the frequency of cells with 4C nuclei reached 50%.     

Industrial process proteomics: alfalfa protein patterns during wet fractionation processing

A proteome reference map of major soluble proteins from Medicago sativa (alfalfa) leaves and stems has been established for the first time. Among 195 spots analyzed by mass spectrometry and N-terminal Edman sequencing, 117 spots were unambiguously identified, representing 87 different proteins. Of these 87 proteins, 13 proteins were directly identified from the partial genome of Medicago sativa, 30 from expressed sequenced tags (ESTs) of the model legume Medicago truncatula and 44 from closely relative species by a cross-species protein identification method. The proteome map of Medicago sativa was then set as a reference to study the major high protein content products that are generated during the wet fractionation process of alfalfa green biomass. Using two-dimensional electrophoresis, we studied the variation of the protein patterns at different steps of the industrial-scale process. We clearly show that the process induces significant changes including chemical modifications, proteolytic events, and heat-shock protein responses. Strikingly, a certain level of cellular regulation is conserved during biomass processing, as exemplified by the induction of some heat shock proteins. Finally, all the results obtained in this proteomic study may help to identify novel products and to improve process designs in alfalfa biomass plants.     

Characterization of Rhizobia from Ineffective Alfalfa Nodules: Ability to Nodulate Bean Plants [Phaseolus vulgaris (L.) Savi.

This study was initiated to characterize Rhizobium isolates obtained from root nodules of ineffectively nodulated, field-grown alfalfa (Medicago sativa L.) plants. The purpose was to determine if these isolates possessed characteristics which would explain either their ineffectiveness in N(2) fixation or their apparent ability to tolerate the moderately acid soil conditions from which they originated. Isolates were characterized by analysis of growth rate, 39 degrees C tolerance, acid production on conventional media, and symbiotic performance. All isolates were ineffective in N(2) fixation on alfalfa, and they contained one or more anomalous characteristics. These included either slow growth rate, lack of 39 degrees C tolerance, or lack of acid production on conventional media. Infectiveness tests on a broad range of legumes revealed that the isolates formed root nodules on M. sativa, Medicago lupulina L., and Phaseolus vulgaris (L.) Savi. (common bean). These results provide evidence that, in some situations, ineffective nodulation of M. sativa in the field may be due to the presence of promiscuous, native Rhizobium species.     

‘陇东’紫花苜蓿原生质体最佳分离条件

以‘陇东’紫花苜蓿（Medicago sativa L.cv.‘Longdong’）下胚轴愈伤组织为材料,研究酶液组合、酶解时间、酶液渗透压、愈伤组织继代培养时间及预处理措施对原生质体分离效果的影响。结果表明：获得产量最高（8.8×105个·g-1）、活力最高（88.55%）的原生质体最佳酶液组合为2%纤维素酶＋0.5%果胶酶＋0.3%崩溃酶、酶解时间为10h、酶液渗透压即甘露醇浓度为0.55mol·L-1,愈伤继代培养天数为12d、预处理措施为4℃、黑暗条件下放置24h。