Influences of Temperature Regime on Germination of Seed of Wild Soybean(Glycine soja)

As an important plant species with high protein contents,wild soybean(Glycine soja),has drawn much attention and appeared to be useful for the genetic improvement of soybean germplasms.Since temperature is one of the numerous environmental factors affecting the germination of most plants,an experimental study was carried out to determine the effect of temperature on germination of wild soybean(G.soja)seeds.Germination test was conducted by setting up thirty-six constant and alternating temperature regimes,ranging from 5 to 40 ℃(16 h night/8 h day).Responses in germination rate to these temperature regimes were then used to construct a quadratic response surface,giving estimated germination rates with confidence intervals at P ≤ 0.05.The results showed that germination capacity was significantly greater while exposed to constant temperatures of 25 ℃,and under the alternating temperature regime the optimum temperature occurred at the 20/25,25/25,25/30 ℃ regime(16 h/8 h)with the amplitude widened from 0 to 5 ℃.Together with regional monthly climate data,these results could be used to improve and promote the cultivation of wild soybean(G.soja),making it available to develop the location-specific optimum seeding time and to apply weed-control treatments.

Screening and evaluation for soybean resistance to Phakopsora pachyrhyzi in Glycine soja

Soybean rust caused by Phakopsora pachyrhizy is an important disease of soybean, and yield losses are very common in humid growing regions. Most commercial cultivars are susceptible and the disease is usually managed with fungicides. Resistance breeding is the most effective in controlling the disease. In this study, detached - leaf assay and greenhouse inoculation were used to screen 200 Glycine soja accessions for resistance to Phakopsora pachyrhizi in China. Most of the accessions were susceptible, and W8214 was the only accession that demonstrated RB resistance to Phakopsora pachyrhizy. The further repeated tests confirmed the resistance in W8214.

国家基因库野生大豆(Glycine soja)资源最近十年考察与研究

总结了国家种质库最近10年野生大豆搜集进展和研究概况。我国野生大豆资源在1979-1982年间搜集并保存在国家基因库的为5939份;1996-2000年搜集了600份;2001-2010最近10年补充搜集了全国17个省(市、区)、318个县(市、旗)、930个乡镇(农场)。其中新搜集县市178个,共收集典型野生大豆资源1979份,野生大豆收集样品新增资源33.3%。

中国野生大豆(Glycine soja)遗传资源主要形态、遗传变异和结构

介绍了我国野生大豆遗传资源主要的形态类型、物种内遗传关系和遗传结构。进化的瓶颈不仅发生在由野生大豆到栽培大豆,也以另一种方式"分化瓶颈"出现于同性状的不同表型类型之间。野生大豆种内种子大小类型是否存在遗传分化?野生和半野生大豆的边界在哪?半野生大豆如何产生的?半野生大豆遗传上密切于栽培种还是野生种?百粒重3～4g的小粒半野生大豆与百粒重8.5g以上的特大粒半野生大豆是否有遗传差异?百粒重8.5g以上的特大粒半野生大豆是否属于栽培大豆?野生大豆的种皮色和种子大小哪个更能反映进化程度?栽培大豆基因是否已经渗入到野生大豆?对这些在学术界常年存在的疑问本文介绍了我们的研究答案。我们认为"真"半野生大豆不存在于现在中国半野生资源收集品中;一些野生大豆中的白花、灰毛、无泥膜性状来源于栽培大豆的基因渗透。

野生大豆(Glycine. soja)抗锈鉴定

大豆锈病近年来已发展成为影响大豆生产的世界性病害。筛选抗锈资源是开展抗病育种的基础,从野生大豆中发掘抗锈资源有利于拓宽抗锈育种的遗传基础。采用离体叶片定量接种方法对513份野生大豆(Glycine so-ja)进行抗锈鉴定,得到抗病资源1份,其余均为感病资源。

野生大豆(Glycine soja)YD63和栽培大豆(G. max)ZD19茎秆解剖结构比较

以野生大豆YD63和栽培大豆ZD19为研究对象,通过组织化学方法观察茎秆解剖结构的差异,分析和阐述这些解剖结构与功能和环境适应性间的关系,旨在为大豆抗逆性研究提供解剖学依据。结果表明:1.野生大豆表皮毛和腺毛多于栽培大豆,且角质层厚度、表皮厚度和表皮比例均大于栽培大豆,表皮和外皮层细胞的木质化和木栓化程度也高于栽培大豆;2.野生大豆皮层、韧皮部、木薄壁组织和髓的比例均大于栽培大豆,茎秆机械强度降低,可塑性升高,抗逆性增强;3.栽培大豆木质部、木纤维和总纤维比例均大于野生大豆,并且表皮细胞壁、韧皮纤维壁、木纤维壁和导管壁厚度均大于野生大豆。栽培大豆组织木质化的比例大于野生大豆,茎秆的机械强度升高,可以更好地维持直立生长和形态构建;4.栽培大豆微管形成层的细胞层数和厚度均大于野生大豆。栽培大豆木质部的比例大于韧皮部的比例,而野生大豆两者比例基本相同;5.野生大豆韧皮部厚壁组织几乎是连续分布,仅在髓射线处中断,而栽培大豆是不连续的,呈片状分布,野生大豆韧皮部厚壁组织的比例大于栽培大豆;6.野生大豆导管壁强度(t/b)2和小导管比例大于栽培大豆,水分运输的安全性较高,但野生大豆木质部的连通性和水分运输的效率低于栽培大豆。

冀东野生大豆(Glycine soja)耐盐碱性鉴定及耐性生理指标测定

为充分利用盐碱地资源,明确野生大豆耐盐碱生理机制,本研究对采集于冀东地区的349份野生大豆种质资源进行耐盐碱性鉴定,测定了高耐材料2010-12和敏感材料2012-34、2012-49在0、100和200 mmol·L^-1盐碱胁迫下的丙二醛、脯氨酸和可溶性糖含量、电解质外渗率以及脯氨酸代谢关键酶吡咯琳-5-羧酸合成酶(GmP5CS)、吡咯琳-5-羧酸还原酶(GmP5CR)、脯氨酸脱氢酶(GmPDH)及谷胱甘肽S-转移酶19(GsGST19)相关基因的表达量。结果表明,表现耐性等级1级的高耐材料有2份(Yong 2和2010-12),表现耐性等级2级的耐性材料有22份,表现耐性等级3、4、5级的材料各有80、77和161份。与CK相比,高耐野生大豆材料2010-12在盐碱胁迫下脯氨酸及可溶性糖含量显著升高,丙二醛含量及电解质外渗率无显著差异。在高耐盐碱野生大豆材料中,GmP5CS、GmP5CR和GsGST19基因表达量上调,GmPDH基因表达量下调。以上结果表明高耐野生大豆材料在盐碱胁迫下脯氨酸合成通路激活,脯氨酸含量升高,说明脯氨酸在野生大豆对抗盐碱胁迫中发挥重要作用。本研究筛选的高耐盐碱野生大豆材料可为培育耐盐碱栽培大豆提供优异种质。

野生大豆(Glycine soja)热击蛋白(HSP)的研究 Ⅱ.松花江下游和长江下游地区野生大豆HSP诱导合成的比较

本文对松花江下游地区野生大豆“０１－３２”和长江下游地区野生大豆“２０１２２”的发育胚根在４０℃条件下进行２，４，６，８和１０ｈｒ的热击处理，分析比较其热击蛋白（ＨＳＰ）诱导合成的种类、累积量及累积动态。研究结果表明：在ＨＳＰ合成种类上，除热击２ｈｒ为松花江下游野生大豆多于长江下游野生大豆，其它热击时间均为后者多于前者；小分子量蛋白的种类，在各热击时间均为长江下游野生大豆多于松花江下游野生大豆。关于ＨＳＰ的累积量，除热击６ｈｒ长江下游野生大豆略低于松花江下游野生大豆，其它热击时间均为前者明显高于后者。

Differentially Expressed Genes of Soybean During Infection by Phytophthora sojae

To elucidate the differential gene expression patterns in soybeans during infection by Phytophthora sojae,a cDNA library for suppression subtractive hybridization （SSH） was constructed with cDNAs from soybean cultivar Suinong 10 treated with sterile distilled water as the driver and cDNAs from Suinong 10 inoculated with P.sojae as the tester.A total of 2 067 recombinant colonies from the SSH library were randomly picked,amplified,and sequenced.After discarding 312 poor quality expressed sequence tags （EST）,1 755 high quality ESTs were assembled and edited to 1 384 tentatively unique genes （TUG）,in which,586 showed significant homology to known sequences,and 798 had low homology or no match with the known sequences.A cDNA microarray containing 307 singletons from the 586 TUGs and 222 singletons from the 798 TUGs was developed to characterize differentially expressed cDNAs in the SSH library,and eight cDNAs were identified to be up-regulated after microarray analysis and then confirmed by real-time PCR.They were homologous to the protein 10,and were also related to some proteins in disease resistance response,such as pathogen-related protein,phenylalanine ammonia-lyase,isoflavone reductase,WRKY transcription factor 31,major allergen Pru ar 1,and pleiotropic drug resistance protein 12.Most of the up-regulated cDNAs encode enzymes of phytoalexin biosynthesis and pathogenesis-related proteins involved in plant disease resistance.Here,we fist reported the Pru ar 1 in soybeans.The findings of this research have contributed to better understanding of soybean resistance to P.sojae at the molecular level.

大豆Soja亚属内种子大小的遗传差异及半野生类型分类归属

Soja亚属包括野生、半野生和栽培大豆三种类型,其中半野生型变异丰富,与野生和栽培大豆形态重叠。然而半野生大豆是属于野生种内还是栽培种的变异类型还存在争议。本研究使用421份包括两组野生大豆百粒重类型、三组半野生大豆百粒重类型和小粒秣食豆类型大豆地方品种对Soja亚属内进行了SSR标记的遗传多样性差异评价和对半野生大豆的分类地位归属问题进行了分析。结果显示,小粒组野生大豆（2.5g以下）和大粒组（2.5~3.0g）有明显的分化。大粒野生大豆和小粒半野生大豆（3.01~5.0g）遗传关系密切,百粒重5.01~10.0g和超过10.0g的半野生大豆组遗传关系密切。大粒野生大豆在遗传上与半野生大豆类型密切。从小粒野生大豆到大粒半野生大豆各类型的遗传分化与它们的百粒重大小有密切关联,即使与小粒秣食豆的百粒重相重叠甚至超过小粒秣食豆的半野生大豆都与小粒栽培大豆有显著的遗传差异。我们所获得的结果清楚地表明：半野生大豆属于野生种内的变异而非栽培种内变异。百粒重大小可以当作评价野生大豆物种内的遗传分化或进化程度的主要指标之一有其遗传上的理论依据。

Establishment of a Core Collection for the Chinese annual wild soybean (Glycine Soja)

With more than 6000 accessions collected from across China, the Chinese National Crop Gene Bank (CNCGB) holds the largest collection of annual wild soybean (Glycine Soja) germplasm in the world. To facilitate the management and utilization of these germplasm collections, it is important to establish a Core Collection. This study compares five sampling strategies, namely random, constant, proportional, logarithmic and genetic diversity-based, to establish a Core Collection for the annual wild soybean germplasm. Among the strategies evaluated, the genetic di- versity-based was found to be the simplest and most efficient. Using hierarchical classification and cluster analysis, in the genetic diversity diversity-based strategy, 652 accessions, accounting for 10.65% of the total 6172 accessions, were se- lected out to represent the total accessions. The established Core Collection has the following features: (i) the 21 de- scriptors observed in the entire collection were compared by the Core Collection, all 18 quality characters of entire collec- tion were preserved by core collection, and coincidence rate of average was 98.4%; (ii) the variant of 13 descriptors of the two collections was very similar, with the coincidence index being 0.96; (iii) the coincidence rate of genetic diversity be- tween the two collections was 81.38% DNA alleles; (iv) the geographic distribution pattern of core collection was the same as the entire collection; (v) molecular marker analysis by 20 SSR primer pairs on 299 accessions showed that the Core Collection covered 83.64% of the entire collection. It thus is concluded that the established Core Collection is rep- resentative and will be a valuable entry point for better evaluation and more efficient utilization of the genetic re- sources available in the annual wild soybean germplasm bank.

A novel TIR-NBS-LRR gene regulates immune response to Phytophthora root rot in soybean

Phytophthora root rot(PRR),caused by Phytophthora sojae,is a devastating disease of soybean.The NBSLRR gene family is a class of plant genes involved in disease resistance.miRNA mediates plant response to biotic stresses by regulating the expression of target genes at the transcriptional or post-translational level.Glyma.16G135500,encoding an NBS-LRR-type protein,is a target of gma-miR1510 that responds to pathogen infections.We cloned and overexpressed Glyma.16G135500(naming it GmTNL16)and knocked down mi R1510 using short tandem target mimic technology to identify the roles of the GmTNL16/gma-mi R1510 pair in the interaction of soybean and the oomycete.By overexpressing GmTNL16 in transgenic hairy roots of soybean,we showed that biomass of P.sojae was lower in overexpressing hairy roots than in control roots.Thus,miR1510 expression was reduced upon P.sojae infection,reflecting the induced expression of GmTNL16 conferring resistance to P.sojae in soybean.Differentially expressed genes were enriched in plant-pathogen interaction,plant hormone signal transduction,and secondary metabolism by RNA sequencing analyze.In particular,jasmonate and salicylic acid pathway-associated genes,including JAZ,COI1,TGA,and PR,responded to P.sojae infection.All of these results indicate that the GmTNL16/gma-miR1510 pair participates in soybean defense response via the JA and SA pathways.

Circadian Rhythm of Root＇s Apical Meristem Mitosis Cells of Soybean

The circadian rhythms of apical meristem mitosis cells of the two types of soybean were investigated： wild soybeans - Glycine max （L.） Merr. （G. hispida Max.）, G. soja Sieb., and Zucc （G, ussuriensis Reg. and Maack）; and the hybrids of their homozygous forms （F6）. Representatives of the initial kinds and interspecific hybrids for reproduction were grown up in comparable conditions. Seeds of a soybean were grown up at 24℃ in Petri cups in thermostat in darkness during winter. For research of mitotic meristem cages activity the tips of roots were fixed temporal by acetoalcohol （fractional fixing） from 1 p.m. till 12 a.m., further up to 12 p.m. each 60 minutes. Acetokarmin was used for preparing the coloring. For researching of mitotic indexes, there have been analyzed from 6 up to 10 thousand cells at each stage （during each moment of time： 01 hour, 02, 03 06 ... 12, 13 ... 18 hours and further）. Within a day （day ＋ night） in cells of root apical meristem, three mitosis ＂waves＂ in each of the investigated forms with an absolute starting point of rest at 12 p.m., with precisely expressed further periodicity were found out. Each of the investigated forms misses in character of wave processes, on a degree of increase and recession of mitosis waves, on the level of mitotic indexes. Maximal mitotic activity at all the investigated forms is noted at 6 and 12 a.m. and 6 p.m.

Inheritance of the Anatomy--Morphological Structure of the Stalk by Interspecific Hybrids of the Glycine L.

The research focuses on the study of anatomical and morphological stalk structure ofsoya interspecific hybrids of the third generation （F3） between （Glycine max （L.） Merr.） and G. soja Sieb. et Zucc. in comparison with parent plant species. The parent plant species and interspecific hybrids were sowed and grew under similar conditions. The similarity of the anatomic structure of stalks of cultivated plants and Glycine soja （wild soya） proves the hypothesis the studied species have the same origin. However, the obtained results show the considerable degree of phylogenetic dissociation between the studied soya species. Interspecific hybrids inherit from G. soja the ability to high intensive growth. The G. soja use in practical selective breeding is of great interest.

野生大豆GsEXPA3基因的生物信息学、逆境转录及毛状根转化分析

为进一步发掘野生大豆中的优质抗逆基因以用于大豆的种质改良,本研究选取积极参与调控植物生长和非生物胁迫抗性的扩展蛋白基因为对象,在野生大豆中克隆了GsEXPA3基因并对其进行了生物信息学分析,通过qRT-PCR检测该基因在逆境胁迫下的转录情况,采用大豆毛状根转化试验分析其对于根系生长的调控作用。结果显示:启动子分析表明GsEXPA3基因的转录可能参与光、脱落酸及干旱胁迫响应并与类黄酮物质合成相关。在低温和干旱胁迫下,GsEXPA3基因的转录呈显著增加趋势,分别在处理后12和9 h达到最高点,为对照组的3.27倍和2.09倍。GsEXPA3基因的过表达显著促进了转基因大豆毛状根的生长,毛状根数量、总根长和总根重,与对照组相比分别提高了56.83%、53.19%和53.35%。结果说明野生大豆扩展蛋白基因GsEXPA3对于栽培大豆的分子育种具有良好的应用价值。

中国栽培和野生大豆农艺品质性状与SSR标记的关联分析 I.群体结构及关联标记

关联作图是一种利用连锁不平衡(linkage disequilibrium,LD)检测自然群体中基因位点及其等位变异的方法。利用60个SSR标记,对全国大豆地方品种群体(393份代表性材料)和野生大豆群体(196份代表性材料)的基因组变异进行扫描,分析两类群体的连锁不平衡位点、群体结构,并采用TASSEL软件的GLM(general linear model)方法对16个农艺、品质性状观测值进行标记与性状的关联分析。结果表明:(1)在公共图谱上不论共线性的或是非共线性的SSR位点组合都有一定程度的LD,说明历史上发生过连锁群间的重组;栽培群体的连锁不平衡成对位点数较野生群体多,但野生群体位点间连锁不平衡程度高,随距离的衰减慢。(2)群体SSR数据遗传结构分析发现,栽培群体和野生群体分别由9和4个亚群体组成,亚群的划分与群体地理生态类型相关联,证实地理生态类型划分有其遗传基础。(3)栽培群体中累计有27个位点与性状相关;野生大豆种质中累计有34个位点与性状相关。部分标记在两类群体中都表现与同一性状关联,检出的位点有一致性,也有互补性;一些标记同时与2个或多个性状相关联,可能是性状相关乃至一因多效的遗传基础;关联位点中累计有24位点(次)与遗传群体连锁分析定位的QTL一致。

不同尺度下野大豆种群的遗传分化

为了阐明不同尺度范围内野大豆种群的遗传分化情况 ,应用随机扩增多态性 DNA( RAPD)方法 ,分别对我国 5个纬度 8个不同地点的野大豆 ( Glycine soja)种群及浙江金华地区 5个野大豆种群 ,进行了分子生态学研究。根据 RAPD数据计算相似系数及遗传距离并进行聚类分析 ,发现无论是不同纬度野大豆种群还是金华地区野大豆小种群均存在较高的遗传变异 ,且不同纬度野大豆种群间的遗传变异与地理纬度有一定正相关。在对金华地区野大豆种群遗传多样性的研究中 ,利用 Shannon指数估算了 5个野大豆种群的遗传多样性 ,发现大部分的遗传变异存在于野大豆种群间 ( 78.5 %) ,只有少部分的遗传变异存在于种群内。本文就此探讨了不同尺度下野大豆种群的遗传多样性与环境因子的关系 ,并对其成因及维持机制进行了讨论。

中国栽培和野生大豆农艺及品质性状与SSR标记的关联分析--Ⅱ．优异等位变异的发掘

在前文研究已检出与农艺品质性状显著关联的SSR位点的基础上,本文进一步对与性状关联位点的等位变异作解析,通过将携带某等位变异的所有材料表型均值与携带无效等位基因(null allele)材料表型均值做比较,估计等位变异的潜在表型效应增量(减量),进一步利用该信息估计位点增效(减效)等位变异的平均效应,鉴别出一批农艺品质性状优异位点、等位变异及携带优异等位变异的载体材料。发现在栽培及野生种质中检出的优异等位变异有同、有异、有互补性。发现关联位点正、负效应等位变异均值间有差异,可根据育种目标性状选择要求,选取适合的位点及相应等位变异。同一标记位点可与多性状关联,其等位变异在不同性状间各有其表型效应的方向和大小;等位变异在相关性状效应上方向、大小的异同解释了性状间正、负相关的遗传原因。关联作图得到的信息可以弥补家系连锁法QTL定位信息的不足,并直接利用等位变异信息进行亲本选拔、组合选配及后代等位条带辅助选择以提高育种成效。

中国栽培大豆和野生大豆不同生态类型群体间遗传演化关系的研究

对全国栽培大豆不同地理、季节生态类型和一年生野生大豆不同地理生态类型的大量材料 ,进行形态、等位酶和细胞器 DNA RFL P性状等不受人工选择直接影响的中性性状的分析。结果表明 :( 1)我国栽培大豆不同生态类型群体间地理生态分化明显 ,遗传距离较大 ;同一地理群体内存在季节生态分化 ,遗传距离相对较小 ;南方春豆与东北春豆熟期组相近 ,但其遗传距离大于与南方其他生态型的距离。 ( 2 )中国栽培大豆在地理上和季节上表现由晚熟型向早熟型进化的趋势 ,东北、黄淮栽培大豆各种生态型与本地区野生大豆群体的遗传距离大于与南方野生大豆群体的遗传距离 ,因而南方原始野生大豆可能是各地栽培大豆的共同祖先 ,南方原始栽培类型在向北方扩展过程中不断演化出各种早熟类型。

野生大豆、半野生大豆和栽培大豆对苗期干旱胁迫的生理反应

以野生、半野生和栽培大豆为材料,在旱棚盆栽条件下进行苗期干旱胁迫试验。对叶绿素含量、光合速率、脯氨酸含量、丙二醛(MDA)含量、相对电导率、可溶性糖(WSS)含量、超氧化物歧化酶(SOD)活性、过氧化物酶(POD)活性、过氧化氢酶(CAT)活性进行测定。结果表明,在干旱胁迫下,叶绿素含量、光合速率、可溶性糖含量、脯氨酸含量及SOD活性表现为野生大豆>半野生大豆>栽培大豆;而丙二醛含量、相对电导率、POD活性则表现为野生大豆<半野生大豆<栽培大豆。干旱胁迫下不同类型材料各生理指标与正常供水条件相比都发生不同程度的变化,叶绿素含量、光合速率降幅为野生大豆<半野生大豆<栽培大豆;脯氨酸含量增幅为野生大豆>半野生大豆>栽培大豆;而丙二醛(MDA)含量、相对电导率、SOD活性、POD活性、CAT活性增幅变化顺序为野生大豆<半野生大豆<栽培大豆。在干旱胁迫下,野生大豆、半野生大豆和栽培大豆的生理指标发生显著变化,3种类型大豆的变化幅度差异显著。野生大豆在干旱胁迫下生理指标的表现优于半野生大豆和栽培大豆。