扁荚山黧豆对NaCl胁迫的形态和生理响应

为了分析土壤盐渍化对扁荚山黧豆(Lathyrus cicera L.)生长发育的影响及其耐盐的生理机制,利用梯度浓度NaCl对扁荚山黧豆种子的萌发期及幼苗期进行了胁迫处理,统计幼苗形态对NaCl胁迫的响应,检测SOD等抗氧化酶活性及脯氨酸等渗透调节物质含量。结果表明:不同浓度的NaCl均可限制扁荚山黧豆的种子萌发率和幼苗生长,其耐受NaCl的浓度阈值介于100~150 mmol/L之间;APX、POD、PPO和SOD是扁荚山黧豆应对NaCl胁迫的主要抗氧化酶类,使活性氧的积累与清除处于稳态;在100 mmol/L NaCl处理条件下,地上部的脯氨酸可能向根部进行了转运;通过地上部与根部的协同作用,增加扁荚山黧豆的盐耐受性。POD、SOD等抗氧化酶活性和脯氨酸、MDA含量等7个生理指标之间呈显著相关性,共同响应NaCl胁迫并维持了扁荚山黧豆的正常生长。

山黧豆AAE超家族基因鉴定及LsAAE3的酶活检测

【目的】三七素(β-N-草酰-L-α,β-二氨基丙酸,β-ODAP)是山黧豆、三七等物种中的神经活性物质,其生物合成与草酰辅酶A密切相关。研究基于全基因组鉴定山黧豆草酰辅酶A合成酶的编码基因,为β-ODAP生物合成的调控研究奠定基础。【方法】用AMP结合结构域隐马尔科夫模型在山黧豆基因组中鉴定酰基激活酶超家族(acyl-activating enzyme superfamily,AAE)基因,在此基础上,克隆山黧豆草酰辅酶A合成酶基因LsAAE3,用多序列比对、原核表达及酶活检测等方法分析LsAAE3基因功能。【结果】山黧豆中至少存在22条AAE家族基因;系统发育分析表明:LsAAE3与拟南芥AtAAE3等聚为一支,隶属于酰基辅酶A合成酶家族。基因克隆及序列分析表明:LsAAE3基因cDNA全长为1566 bp;LsAAE3和AtAAE3的保守结构域类似,均含有AMP结合位点、CoA结合位点及AAE保守结构域。SDS-PAGE检测表明所获融合蛋白条带单一,大小在56 kD左右;用His标签抗体进行Western blot分析发现,诱导后和纯化的融合蛋白均能检测到特征条带,说明所获融合蛋白为LsAAE3。体外酶活检测表明:LsAAE3具有草酰辅酶A合成酶活性,其活性发挥依赖于ATP、Mg^(2+)。【结论】研究在全基因组水平鉴定山黧豆AAE超家族基因,并验证了LsAAE3具有草酰辅酶A合成酶活性,为进一步分析山黧豆LsAAE3在β-ODAP生物合成过程中的功能奠定基础。

山黧豆还田与氮肥减施对稻田土壤活性有机碳组分及酶活性的影响

为探明绿肥山黧豆(Lathyrus sativus)还田配施氮肥在改良土壤方面的效应,明确合适的绿肥配施氮肥比例,设置2×4双因素试验,研究绿肥的不同翻压量[15000(M_(1))、22500(M_(2))、30000(M_(3))、37500(M_(4))kg·hm^(−2)]和氮肥的不同施氮量[常规施氮量的60%(N_(1))和80%(N_(2))配比]对土壤活性有机碳库各组分、碳库管理指数和酶活性等指标的影响。结果表明:与常规施肥(CF)处理相比,翻压一定量的绿肥并配施减量氮肥能有效提升稻田总有机碳、活性有机碳、可溶性有机碳、微生物生物量碳含量及碳库管理指数,提升效果随配施比例的不同存在差异,其中M_(4)N_(1)、M_(4)N_(2)处理提升效果最佳。在相同施氮水平下,有机碳各组分含量、碳库管理指数及总体酶活性均呈现出随翻压量增加而增加的趋势。与CF处理相比,翻压绿肥并配施氮肥对稻田土壤过氧化氢酶无显著影响(P>0.05),对纤维素酶、蔗糖酶和β-葡萄糖甘酶活性均具有显著影响(P<0.05)。总体酶活性均表现为M_(4)N_(2)>M_(4)N_(1)>M_(3)N_(1)>M_(3)N_(2)>M_(2)N_(1)>M_(2)N_(2)>M_(1)N_(1)>M_(1)N_(2)>CF>CK处理。各有机碳组分之间具有显著(P<0.05)或极显著(P<0.01)的相关性,β-葡萄糖甘酶、纤维素酶与土壤活性有机碳各组分均呈正相关关系(P<0.05)。对土壤活性有机碳组分含量及土壤酶活性影响因素的灰色关联度综合分析结果表明,60%氮肥+37500 kg·hm^(−2)绿肥模式的综合评价效果最好。

Determination of Neurotoxin β-ODAP and Non-protein Amino Acids in Lathyrus Sativus by High-Performance Liquid Chromatography with Precolumn Derivatization with 6-Amino quinolyl-N-hydroxysuccinimidyl Carbamate (AQC)

A new method was developed for the quantitative determination of the neurotoxic nonprotein amino acid, 3-N-oxalyl-L-2,3-diaminopropioni acid (β-ODAP), its nontoxic α-isomer and other non-protein amino acids in the plant samples of Lathyrus sativus after derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) by reversed-phase high-performance liquid chromatography (HPLC). 2-Amino butyric acid (ABA) was used as an internal standard.The RP HPLC detection limit for both isomers is 1.8 ng with good response linearity. The results are compared with a colorimetric method.

Population structure analysis and determination of neurotoxin content in a set of grass pea (Lathyrus sativus L.) accessions of Bangladesh origin

Grass pea(Lathyrus sativus L.) is a crop that is considered one of the more resilient to climate change. With protein-rich seeds and leaves, it has strong potential as human food as well as animal feed and fodder. However, genetic improvement in this crop remains stagnant owing to the poor characterization of its genetic resources. In this study, we characterized 118 accessions of grass pea with 18 EST-SSR markers. A total of 118 accessions, 101 of L. sativus(100 cultivated accessions from Bangladesh and one wild accession) and 17 wild accessions of other Lathyrus species, were used. A total of 67 alleles were detected, with an average of 3.72 alleles per locus and average polymorphism information content of 0.52. A dissimilarity matrix was formed and hierarchical cluster analysis performed using the UPGMA method grouped genotypes into four main clusters. Cluster analysis based on the genetic dissimilarity revealed a clear grouping of the 100 cultivated and 18 wild accessions into four main groups. Group I consisted of 20 accessions with high β-N-oxalyl-L-α,β-diaminopropionic acid(β-ODAP) concentration. Of these 20 accessions, 17 were wild accessions. Only one wild accession(L. cicera) was clustered in group II, which contained 35 accessions. Most of the group II accessions contained low β-ODAP. Group III was represented by 34 accessions, many of them with high β-ODAP. Group IV consisted of 29 accessions, a few of which had very high β-ODAP concentrations. Analysis of molecular variance of the microsatellite data showed significantly higher values of molecular variance between(83%) than within(17%) populations. These characterized accessions will be useful in grass pea breeding programs.

HPLC Determination of Neurotoxin β-N-oxalyl-L-α, β-diaminopropionic acid and Its α -Isomer in Lathyrus sativus by Precolumn Derivatisation with 1-Fluoro-2,4-dinitrobenzcne

A rapid and simple method is presented for determining β-N-oxalyl-α. β- diaminopropionic acid (β -ODAP) and its much less toxic α -isomer (α -ODAP) in Lathyrus sativus. Seed and foliage extracts of Lathyrus sativus were treated with 1-fluoro-2,4-dinitrobenzene (FDNB) and a reversed-phase high-performance liquid chromatographic method for the separation of the derivatives in the pmol range is reported.

海边香豌豆（Lathyrus maritimus）资源开发基础研究

海边香豌豆是生长在青岛海边的野生豆科植物，具有开发前途和应用价值。研究表明，它具有耐盐性、多年生、再生力强、抗逆性强、营养丰富等特点。其核型为Ｋ（２ｎ）＝１４＝６ｍ＋８ｓｍ。

Effect of Dietary Administration of Lathyrus sativus Pulse on Intestinal Biochemical Parameters in Normal and Scorbutic Guinea Pigs

Objective In order to investigate that ascorbic acid deficiency is responcible for lathyrus toxicity, the effect of dietary feeding of lathyrus pulse in normal and scorbutic guinea pigs for 3 months, on intestinal biochemical parameters was undertaken. Methods The intestinal brush border membrane (BBM) marker and xenobiotic metabolising enzymes (XME) were assayed. Results Exposure to 80% lathyrus alone and in scorbutic conditions showed significant inhibition of alkaline phosphatase (28%-30%), sucrase (19%) and γ-glutamyl transpeptidase (GGT) (15%-27%) enzymes, while Ca2+-Mg2+-ATPase was significantly inhibited (38%) in scorbutic plus lathyrus treated group. The phase I XME (AHH) remained unchanged while the phase II enzyme glutathione-S-tranferase (GST) was significantly decreased (20%-22%) in lathyrus and scorbutic plus lathyrus treated groups. Quinone reductase (QR) activity was found to be significantly decreased in lathyrus exposed group (20%). The intestinal biomarker contents including hexose (25%-34%) and phospholipids (20%-40%) were significantly reduced in lathyrus and scorbutic plus lathyrus exposed animals, while sialic acid showed a significant decrease (28%) in scorbutic plus lathyrus treated group. However, cholesterol levels were signifcantly enhanced (15%-28%) in lathyrus and scorbutic plus lathyrus treated animals. Conclusion The results indicate that oral feeding of lathyrus pulse to guinea pigs can alter BBM parameters as well as XME, which may result in the intestinal toxicity. Further, ascorbic acid deficiency could be one of the pre-disposing factors of lathyrus toxicity.

L-Homoarginine Accumulation in Grass Pea (<i>Lathyrus sativus</i>L.) Dry Seeds. A Preliminary Survey

Grass pea (Lathyrus sativus L.) has great agronomic potential as grain and forage legume, and presently is con-sidered as a model crop for sustainable agriculture. However, the development into an important food legume has been hin- dered by the presence of the neurotoxic amino acid β-N-Oxalyl-α, β-diaminopropionic acid (β-ODAP). Recent studies reported that homoarginine (Har) can counteract this toxic action. This research was undertaken to shed light on the variation of Har amount within grass pea. The influence of the environment and of the year-to-year variation of cli- matic conditions was also investigated. Seven Italian grass pea ecotypes were evaluated for two subsequent growing seasons in two locations of southern Italy. In contrast with previous studies collected data evidenced a significant variation of Har amount among the tested ecotypes. Moreover, a significant positive correlation between Har and ODAP level was observed. The effect of year-to-year variation of temperature and rainfall quantity is also discussed.

Effects of Supplementing Finger Millet (<i>Eleucine coracana</i>) Straw with Grass Pea (<i>Lathyrus sativus</i>L.) Hulls and Concentrate Mixture on Feed Intake, Digestibility and Body Weight Change in Washera Sheep

The productivity of sheep in Ethiopia is very low mainly due to a serious shortage of feedstuff. As a result, it is necessary to look for cheap locally available feedstuffs to sustain sheep production in the country. Non-conventional feeds like grass pea (Lathyrus sativus L.) hulls (GPH) could partly fill the gap in the feed supply, decrease competition for food between humans and animals and reduce feed cost. Therefore, this experiment was designed to evaluate effects of different proportions of grass pea hulls and concentrate mixture (CM) supplementation on feed intake, digestibility, and growth of Washera sheep fed finger millet straw (FMS) as a basal diet. Diets consisted FMS alone fed ad libitum (T1), 100% CM (T2), 30% GPH mixed with 70% CM (T3), 30% CM mixed with 70% GPH (T4), and 100% GPH (T5). For all treatments, FMS was fed ad libitum. Experimental sheep were arranged in a randomized complete block design and treatment feeds were assigned to the animals within a block. The results showed that FMS contained 5.5% CP and 68.9% NDF. The supplements contained 21%, 20%, 16% and 14% CP;and 36%, 43%, 52% and 59% NDF for T2, T3, T4 and T5, respectively. Supplementation improved (p < 0.0001) DM and nutrient digestibility. Average daily gain (ADG) was 60, 84, 51, and 28 g/day for T2, T3, T4 and T5, respectively, which showed that T3 recorded the highest ADG and T5 resulted in the lowest ADG (p < 0.0001). While, animals in T1 lost a bodyweight of 5.6 g/day. As the proportion of GPH supplementation was greater than 30%, there was reduction (p < 0.05) in DM intake, DM and CP digestibility and consequently depressed ADG in sheep. Partial budget analysis results showed that net return and change in net return were in the order of T3 > T2 > T4 > T5. Therefore, from biological point of view as well as based on partial budget analysis, T3 was found to be recommendable for smallholder sheep producers.

绿肥山黧豆在四川省的生产与应用现状

山黧豆营养丰富、适应性强、生物产量高、分布广泛,是不可多得的肥、饲、粮作物,具有较好的经济、社会、生态效益。在新形势下,农业产业结构不断调整,绿肥作为有机肥源作用日益突出。本文综述了四川省绿肥山黧豆的发展历程、高产栽培和利用技术、生产应用模式、应用前景,以期将其应用于现代农业生产中,对进一步加快四川省生态文明建设和助力乡村振兴有着积极意义。

不同种类绿肥翻压对土壤肥力的影响

通过模拟试验,研究了旱地翻压箭筈豌豆(Vicia sativa L.)、苕子(Vicia villosa Roth.Var.)和山黧豆(Lathyruspalustris L.Var.Pilosus ledeb)三种绿肥对土壤肥力特性的影响。试验结果表明,在绿肥翻压腐解过程中,土壤pH(原始pH值为5.95)均呈现出先升高后降低最后趋于平缓的趋势,在翻压0～25 d内高于不翻压绿肥对照,之后始终低于对照。翻压绿肥处理的土壤有机碳含量均高于对照,增幅为1.6%～6.2%。土壤无机氮含量显著高于对照,无机氮形态0～20 d以铵态氮为主,30～70 d以硝态氮为主,翻压70 d时,硝态氮含量较对照增加了15倍以上。翻压绿肥后土壤速效磷的含量均高于对照,增幅为11.9%～37.7%。翻压绿肥的土壤速效钾含量在翻压15 d时达最高且均比对照增加1.5倍以上。不同绿肥对土壤养分特性的影响不同,山黧豆处理的土壤pH变化幅度最大,土壤有机碳和无机氮含量最高,箭筈豌豆处理的土壤速效钾和速效磷含量最高。本试验条件下,三种绿肥翻压均能够改变土壤pH值,提高土壤硝态氮和速效钾含量,土壤有机碳和速效磷含量略有增加。

高山草原条件下一年生豆科牧草生产性能的评价

1998年对自国际干旱农业研究中心 (ICARDA)引进的野豌豆属 (Viciaspp .)和山黧豆属(Lathyrusspp .)的 5个种 50个品系在高山草原条件下的牧草和种子产量表现作了评价 ,1999年对其中能够完成生育期的春箭舌豌豆 9个品系、法国野豌豆 4个品系、鹰嘴山黧豆 2个品系和栽培山黧豆 1个品系做了进一步的测定。这 16个品系 1998年和 1999年的平均产草量分别是春箭舌豌豆 6 50 9.5和 5489.1kg/hm2 ,法国野豌豆 3398.0和 2 786 .7kg/hm2 ,鹰嘴山黧豆 76 10 .3和 476 1.9kg/hm2 ,栽培山黧豆 580 4 .7和 3858.2kg/hm2 ;种子产量分别是春箭舌豌豆 50 9.7和 10 10 .0kg/hm2 ,法国野豌豆 536 .0和 10 73.6kg/hm2 ,鹰嘴山黧豆 152 .2和 732 .7kg/hm2 ,栽培山黧豆 16 .5和 12 .8kg/hm2 。长柔毛野豌豆的所有 10个品系、栽培山黧豆的 9个品系和鹰嘴山黧豆的 8个品系在当地不能完成生育期。根据两年的试验结果 ,从牧草和种子产量两方面综合评价 ,春箭舌豌豆更适合在高山草原种植。春箭舌豌豆 2 556、2 56 0和 2 56 6等品系地上部分生物量与对照品种 333/A狭叶野豌豆相当 ,但种子产量比对照种高 19.6 %～ 77.4 % ,表现出较高的生产性能和适应性 ,值得进一步选育。春箭舌豌豆品系的牧草与种子产量呈负相关关系 ,在品种选育过?

水分胁迫下山黧豆中ABA及ODAP的积累研究

用PEG、PEG +ABA、ABA分别处理 15d龄的山黧豆幼苗 ,取其叶片为实验材料 ,测定内源ABA、O DAP、MDA和H2 O2 含量以及几种抗氧化酶活性 ,结果表明 ,与对照相比处理材料叶片中ABA和ODAP含量显著增加 ;外源ABA的加入降低了PEG胁迫引起的MDA和H2 O2 含量的增加 ,延缓了PEG胁迫引起的CAT活性的衰减 ,提高了GR活性 .用外源ABA长时间处理山黧豆 ,发现叶片中ABA含量显著增加 ,随后出现ODAP的积累 ;ABA处理初期 (0～ 3d)对叶片中活性氧代谢影响不大 ,随着ABA处理时间的延长 (7～ 15d) ,可引起叶片中SOD、POD、CAT、GR活性的降低 ,MDA、H2 O2 含量的增加 ,表明ABA确实可促进ODAP的积累 .

山黧豆毒素ODAP的生物合成及与抗逆性关系研究进展

有关山黧豆毒素ODAP的生物合成途径的前体物和合成程序已经证实,但其关键合成酶尚未分离与鉴定,因而无法克隆相应基因和利用反义RNA技术以控制其生物合成。研究表明,利用相应的抑制剂控制ODAP生物合成前体物可降低ODAP的积累量。山黧豆中ODAP含量与其抗逆性之间密切相关,这与其能有效地清除山黧豆中.OH自由基有关,外源ODAP处理获同样效果。此外,因ODAP既是含氮化合物,又是游离氨基酸,极易溶于水,可在逆境胁迫下与脯氨酸和多胺一样大量而迅速地积累,推测它也可能作为细胞渗透调节物质和防脱水剂,并在氮代谢和能量代谢方面起重要作用。对今后该领域的重点研究方向也进行了探讨。

山黧豆低毒品种的筛选及栽培利用技术研究

对国内外引进的６５个山黧豆品种，经３年观察和农艺性状的测定以及β－草酰氨基丙氨酸（ＢＯＡＡ）、营养成分的分析，筛选出产量、蛋白质含量高，毒素含量低，经毒理学实验为安全饲用的黑龙江山黧豆－１、黑龙江山黧豆－２、西德山黧豆３个品种供生产中应用。还针对山黧豆作为饲草（料）推广应用中的播种量、磷肥施用量、压青及根茬肥效等栽培利用措施进行了探讨。

山黧豆叶片蛋白质双向电泳技术的建立

以山黧豆叶片为材料,比较分析了蛋白质的不同提取方法,在此基础上着重于样品制备.对IPG胶条的选择,第一向等电聚焦和第二向SDS-聚丙烯酰胺凝胶电泳的电泳程序及参数、染色方法等相关技术进行了比较和条件优化.结果显示:采用TCA-丙酮沉淀法提取蛋白质,裂解液中加入Tris-base作为蛋白酶抑制剂,等电聚焦电泳时延长低电压的电泳时间(30 V1、2 h,500 V1、h,1 000 V2、h)以促进盐离子泳出的方法对山黧豆叶片蛋白质进行双向电泳,并用考马斯亮蓝和银染复合染色法进行凝胶染色,能够获得蛋白点清晰的双向电泳图谱,说明用优化后的方法建立起的山黧豆叶片蛋白质双向电泳技术,蛋白质样品制备质量好,电泳分辨率高,完全适合于进一步的蛋白质组学研究.

家山黧豆及其毒素ODAP的研究

家山黧豆作为人类的食物及动物的饲料,最早可追溯到新石器时代.长期以来,因其营养丰富,具有耐旱、耐寒等优良生产性状,广受世界各地的青睐.特别是在大旱年份,在其它粮食作物绝收的情况下,仍有较好的收成,因此,被视为干旱及半干旱地区首选的优良作物.但由于其含有毒素β-ODAP,使山黧豆的种植受到限制.近年来,由于人类对肉质品的量与质的多元化的需求,家山黧豆这一潜在的、丰产的、高营养的豆科作物,已引起各国学者的广泛关注.从家山黧豆在植物学、遗传学、分子生物学、生态学、营养学、山黧豆中毒、毒理学、毒素ODAP、ODAP的分析方法、ODAP的生物合成途径、养殖业等几个方面的研究进展作了简要综述.

山黧豆根系对H_2O_2诱导氧化胁迫的生理应答

以水培7d苗龄的山黧豆幼苗为材料,向水培溶液中施加不同浓度H2O2处理山黧豆幼苗24h,分析山黧豆根系受氧化胁迫的程度与抗氧化系统的应答特征,以揭示山黧豆对氧化胁迫的耐受机制。结果显示:(1)随外源H2O2处理浓度的不断增加,山黧豆幼苗侧根的数目无显著变化,而其根的鲜重则显著降低。(2)同时,根系组织的内源H2O2染色范围和程度显著增高,但根尖区域始终保持较低水平的H2O2;相反,O-·2染色范围和程度明显减少,根尖区域却始终保持较高水平的O-·2。(3)同期根系抗坏血酸(ASC)含量及过氧化氢酶(CAT)、过氧化物酶(POD)与抗坏血酸过氧化物酶(APX)的活性均表现出了先升高后降低的趋势,而超氧化物歧化酶(SOD)一直表现为持续上升的趋势。研究表明,在外源H2O2胁迫条件下,山黧豆根系O-·2的积累可能与其生长和活力呈正相关,而根系H2O2的积累则与其受氧化胁迫程度呈正相关;低浓度的H2O2处理可以提高山黧豆抗氧化系统对体内活性氧的清除能力。

水分胁迫对山黧豆萌发中蛋白质和游离氨基酸的影响

对山黧豆 (L athyrus sativus)进行不同浓度 PEG胁迫处理 ,研究水分胁迫对山黧豆种子萌发过程中蛋白质和游离氨基酸的影响。结果表明 ,水分胁迫能导致山黧豆根芽和子叶可溶性蛋白质含量增加 ,非可溶性蛋白质分解受到抑制 ;子叶游离氨基酸含量降低 ,而根芽游离氨基酸含量增高 ,尤其是丙氨酸、异亮氨酸、脯氨酸和精氨酸。随着种子萌发时间的延长 ,根芽 β- ODAP含量逐渐增加 ,而子叶 β- ODAP却降低 ;在水分胁迫下子叶和根芽