基于AFLP标记的中国西藏近缘野生大麦遗传多样性分析

选取7对引物组合, 构建了36份西藏近缘野生大麦和4份栽培大麦的AFLP指纹图谱, 共获得清晰可辨的条带227条, 其中多态性带194条, 占85.46％, 从DNA分子水平显示出所试材料遗传多样性较为丰富。计算得各样品间的遗传距离(欧氏距离)介于2.646~10.488之间, 应用离差平方和法对供试材料的AFLP数据结果进行聚类, 建立了40份大麦材料的AFLP树状图, 聚类结果将40份材料分为5类, 进一步揭示出供试材料间遗传背景的相似性和复杂性。结合Nei’s遗传一致性分析结果, 发现近缘六棱野生大麦较之近缘二棱野生与栽培大麦的亲缘关系更近, 支持栽培大麦是从野生二棱大麦起源, 而野生六棱大麦是进化过程中的过渡类型的大麦系统发生观点。     

从甘孜野生二棱大麦的发现论栽培大麦的起源和种系发生”

1964-1965两年,我们先后在甘孜、道孚两地发现了野生二棱、六棱、中间型大麦三种。1966-1968年,将三种野生大麦分别与栽培六棱裸粒大麦杂交,结果是：二棱、碎穗、有移三种性状都是显性,与其相对性状在F,呈3:1的比例;F：穗型、碎性、祥状都有不同程度的中间类型出现。三种野生型中,只有二棱型是完全纯合的;六棱型有纯合的,也有杂合的;中间型完全是杂合的。由此证明,只有野生二棱大麦才是真正的野生种。因此认为：大麦的六棱、坚穗、裸粒系由二棱、碎穗、有释进化而来,栽培大麦的始祖是Hordes. spoatnneu.o从廿孜野生二棱大麦的发现及对其遗传、生态的分析,可以认为栽培大麦起源于我国西南高原地区。     

应用微卫星标记研究西藏野生大麦的遗传多样性

以西藏不同地区的106份野生大麦为材料,其中包括50份野生二棱大麦（HS）,27份野生瓶形大麦（HL）和29份野生六棱大麦（HA）,用Liu等（1996）发表的SSR连锁图的每个连锁群的两个臂的不同位置上选取3～5个共30个SSR标记,研究了西藏3类野生大麦的遗传多样性。结果表明,这3类野生大麦在遗传组成及等位变异频率分布上存在着明显的遗传分化。在总样本中,共检测到229个等位变异,平均每个SSR位点检测到7．6个等位变异,其中70个为这3类野生大麦间共同的等位变异,等位变异数在这3类野生大麦间有明显的差异,亚种问的遗传多样性明显高于亚种内的遗传多样性。其遗传多样性大小顺序为HS〉HL〉HA。聚类分析表明,野生二棱大麦、野生六棱大麦分别聚在不同的两类,而野生瓶形大麦中各有约50％的材料分别聚在这两类。根据本研究及前人研究结果,我们认为中国栽培大麦是从野生二棱大麦经野生瓶形大麦向野生六棱大麦进化的。该结果支持了栽培大麦起源的“野生二棱大麦单系起源论”的观点。     

青藏高原栽培大麦与近缘野生大麦的核型和酯酶同工酶分析

本文通过对青藏高原栽培大麦与近缘野生大麦的核型和酯酶同工酶分析,研究了它们之间的亲缘关系。核型分析表明,各野生大麦和栽培大麦都是2n=14,x=7,核型都基本相似。但在第1号染色体的臂比上,从二稜野生大麦到六稜野生大麦再到栽培大麦有一个从非等臂到等臂的发展趋势,还有第6号染色体短臂上的随体,栽培大麦显著大于各野生大麦。酯酶同工酶分析表明,栽培大麦的酶谱与六稜野生大麦的很相似,而与二稜野生大麦的差异显著。以上说明,青藏高原近缘野生大麦与栽培大麦的亲缘关系密切,其中六稜野生大麦比二稜野生大麦更接近于栽培大麦。     

青藏高原近缘野生大麦5S rRNA基因染色体原位杂交定位

采用原位杂交技术,以５Ｓ ｒＲＮＡ基因为探针,对产于青藏高原的４份近缘野生大麦和栽培大麦,即：二棱野生大麦Ｈｏｒｄｅｕｍ ｖｕｌｇａｒｅ Ｌ．ｓｓｐ．ｓｐｏｎｔａｎｅｕｍ（Ｋｏｃｈ）Ｈｓｕｅ,六棱野生大麦Ｈ．ｖｕｌｇａｒｅ Ｌ．ｓｓｐ．ａｇｒｉｏｃｒｉｔｈｏｎ（Ａｂｅｒｇ） Ｈｓｕｅ,六棱瓶形野生大麦Ｈ．ｖｕｌｇａｒｅ Ｌ．ｓｓｐ．ａｇｒｉｏｃｒｉｔｈｏｎ ｖａｒ．ｌａｇｕｎｃｕｌｉｆｏｒｍ（Ｂａｋｈｔ） Ｈｓｕｅ,栽培大麦Ｈ．ｖｕｌｇａｒｅ．Ｌ．进行了研究,将杂交结果进行观察与统计,并建立起５Ｓ ｒＲＮＡ基因定位的模式图。结果表明５Ｓ ｒＲＮＡ基因在染色体上的位点呈现动态变化,由二棱野生大麦、六棱瓶形野生大麦到六棱野生大麦、栽培大麦、位点数目有递增的趋势,而且位置也发生了某些改变。探讨了５Ｓ ｒＲＮＡ基因进化     

青藏高原野生大麦和瓦兰大麦的核型及带型研究

本文报道了我国特有大麦——青藏高原野生普通大麦和瓦兰大麦的核型及N带带型。其核型均方2n=2x=14=12m(2SAT)＋2sm(2SAT),均属2A类型。野生普通大麦的N带带型为2n=2x=14=瓦兰大麦的N带带型为2n=2x=14=昌＋2 CI CI＋,昌＋,CI 2CI＋2 CI＋2 CIN CI＋,CI 2 T-1,野生普通大麦、瓦兰大麦与栽培大麦在核型无明显差异,野生普通大麦与瓦兰大麦带型差异明显,且野生普通大麦与栽培大麦间的带) NJ差异大于瓦兰大麦与栽培大麦间的带型差异。分析表明,在进化关系上,野生普通大麦要原始一些。瓦兰大麦要进化一些。文章还对大麦核型及显带等方面的研究进行了讨论。     

美国不同棱型大麦种质资源品质分析

对新引进的300份美国大麦种质资源不同棱型间蛋白质、赖氨酸和淀粉含量的鉴定结果分析表明:①二棱大麦蛋白质总体方差与六棱大麦具有极显著的差异,其频率分布分别是以含量12.5%和17.0%为中心的双峰分布,这可能反映了二棱大麦利用上的新特点;淀粉含量二棱大麦显著高于六棱;六棱大麦当蛋白质作为固定变量时,赖氨酸与淀粉的相关性不显著.②提出了促进SPSS统计分析软件在农业科研上的开发应用研究的建议.     

栽培大麦的起源与进化— 我国西藏和川西的野生大麦

       文报道了在我国西南地区发现有二棱野生大麦。它有广大的分布地区。形态学鉴定,证明它们分属于二棱野生大麦的三个基本变种。还证明六棱野生大麦有广泛的分布地区和物种的多形性。a次证明我国西南地区有六棱瓶形野生大麦,此外还有一些野生大麦的中间类型分布在这一地区。       细胞学鉴定证明上述野生大麦类型染色体组型的第一对最长染色体的一端携带小随体,属a,a,型。       有些野生大麦具有抗病、丰产、优质的特点,可作为育种家的珍贵原始材料。       根据上述实际资料认为二棱野生大麦、瓶形野生大麦和六楼野生大麦是野生大麦由野生向栽培方向进化过程中所经历的几个进化阶段。这些类型全部生长在我国西南地区。说明我国是栽培大麦的起源中心之一。     

青藏高原野生大麦和瓦兰大麦的核型及带型研究

本文报道了我国特有大麦——青藏高原野生普通大麦和瓦兰大麦的核型及N带带型。其核型均为2n=2x=14=12m(2SAT)+2sm(2SAT),均属2A类型。野生普通大麦的N带带型为2n=2x=14=2 C/CI+2 C/CI+2 C/CI+2 CI/CI+2 C/CI+2 CN/CI+2 CI/CI,瓦兰大麦的N带带型为2n=2x=14=2 C/CI+2 C/CI+2 C/CI+2 CI/CI+2 C/CI+2 CIN/CI+2 CI/CI,野生普通大麦、瓦兰大麦与栽培大麦在核型上无明显差异,野生普通大麦与瓦兰大麦带型差异明显,且野生普通大麦与栽培大麦间的带型差异大于瓦兰大麦与栽培大麦间的带型差异。分析表明,在进化关系上,野生普通大麦要原始一些,瓦兰大麦要进化一些。文章还对大麦核型及显带等方面的研究进行了讨论。     

中国近缘野生大麦醇溶蛋白的遗传多态性研究

随机选取来源于中国西藏24个不同行政县的181份近缘野生大科材料,其中包括47份六棱野生大麦,134份二棱野生大麦,选用青藏高原的二棱野生大麦（W2）,欧洲的代表品种Betzes大麦及以色列的二棱野生大麦（Is)作对照。利用A-PAGE（Acidic-polyacrylamide gel electrophoresis)法进行了醇溶蛋白遗传多态性的研究。结果表明,184份供试材料共得到60种不同的电泳图谱,说明西藏近缘野生大麦醇溶蛋白遗传多态性非常丰富,其中有38种图谱为单一材料所独有,以ZYM0019和ZYM1488材料为代表的2种醇溶蛋白图谱占供试材料的29.3%,在西藏分布较为普遍,聚类分析表明地理环境相似的地区有着相似的图谱类型,图谱类型与地理生态环境具有一定的相关性。讨论了中国近缘野生大麦醇溶蛋白的多态性与地理分布的关系。以及栽培大麦的起源中心等问题。     

Hordein variation in the genus Hordeum as recognized by monoclonal antibodies.

The composition of the major storage protein, hordein, in wild barley species has been studied by using gel electrophoresis, Coomassie staining, and immunoblot assays. We have shown earlier that it is possible to obtain cross-reaction outside the cultivated barley, with monoclonal antibodies raised against hordeins from the barley cultivar Bomi. These antibodies have now been used to investigate the hordein composition in all species of the Hordeum genus. The results showed that polypeptides similar to the two major hordein groups of cultivated barley, the B- and C-hordeins, are produced in all wild Hordeum species, and that there are both similarities and differences between the two hordein groups. The similarities indicate a common evolutionary origin, while the distinction between B- and C-hordeins in the entire genus clearly shows that the divergence of their coding genes preceded the divergence of the Hordeum species. The presence of the same antigenic site in two different species indicates that they are evolutionarily related. Among the wild species, two rarely occurring sites were exclusively found in H. vulgare ssp. spontaneum and H. bulbosum, which confirms that they are the cultivated barley's closest relatives. Some of the antibodies also gave an extensive reaction pattern with H. murinum, which suggests a fairly close relationship to H. vulgare, though not as close as between H. vulgare and H. bulbosum.     

Phylogenetic analysis of the genus Hordeum using repetitive DNA sequences

A set of six cloned barley (Hordeum vulgare) repetitive DNA sequences was used for the analysis of phylogenetic relationships among 31 species (46 taxa) of the genus Hordeum, using molecular hybridization techniques. in situ hybridization experiments showed dispersed organization of the sequences over all chromosomes of H. vulgare and the wild barley species H. bulbosum, H. marinum and H. murinum. Southern blot hybridization revealed different levels of polymorphism among barley species and the RFLP data were used to generate a phylogenetic tree for the genus Hordeum. Our data are in a good agreement with the classification system which suggests the division of the genus into four major groups, containing the genomes I, X, Y, and H. However, our investigation also supports previous molecular studies of barley species where the unique position of H. bulbosum has been pointed out. In our experiments, H. bulbosum generally had hybridization patterns different from those of H. vulgare, although both carry the I genome. Based on our results we present a hypothesis concerning the possible origin and phylogeny of the polyploid barley species H. secalinum, H. depressum and the H. brachyantherum complex.     

A barley gene family homologous to the maize rust resistance gene <Emphasis Type="Italic">Rp1-D</Emphasis>

Many characterized plant disease resistance genes encode proteins which have conserved motifs such as the nucleotide binding site. Conservation extends across different species, therefore resistance genes from one species can be used to isolate homologous regions from another by employing DNA sequences encoding conserved protein motifs as probes. Here we report the isolation and characterization of a barley (Hordeum vulgare L.) resistance gene analog family consisting of nine members homologous to the maize rust resistance gene Rp1-D. Five barley Rp1-D homologues are clustered within approximately 400 kb on chromosome 1(7H), near, but not co-segregating with, the barley stem rust resistance gene Rpg1; while others are localized on chromosomes 3(3H), 5(1H), 6(6H) and 7(5H). Analyses of predicted amino-acid sequences of the barley Rp1-D homologues and comparison with known plant disease resistance genes are presented.     

High capacity of plant regeneration from callus of interspecific hybrids with cultivated barley (Hordeum vulgare L.)

Callus was induced from hybrids between cultivated barley (Hordeum vulgare L. ssp. vulgare) and ten species of wild barley (Hordeum L.) as well as from one backcross line ((H. lechleri x H. vulgare) x H. vulgare). Successful callus induction and regeneration of plants were achieved from explants of young spikes on the barley medium J 25–8. The capacity for plant regeneration was dependent on the wild parental species. In particular, combinations with four related wild species, viz. H. jubatum, H. roshevitzii, H. lechleri, and H. procerum, regenerated high numbers of plants from calli.     

Characterization by RFLP analysis and genomic in situ hybridization of a recombinant and a monosomic substitution plant derived from Hordeum vulgare L. x Hordeum bulbosum L. crosses.

Interspecific crosses in Hordeum have been made with the aim of transferring desirable traits, such as disease resistance, from a wild species, Hordeum bulbosum, into cultivated barley (Hordeum vulgare). Interspecific recombinants have previously been identified using several methods, but there are limitations with all the techniques. We improved our ability to characterize progeny from H. vulgare x H. bulbosum crosses by using genomic in situ hybridization (GISH). The plant material comprised a recombinant and a monosomic alien substitution plant derived from H. vulgare x H. bulbosum crosses. The recombinant possesses a pubescent leaf sheath conferred by a gene transferred from H. bulbosum into barley cultivar Golden Promise. The use of GISH on a plant homozygous for the pubescence gene confirmed the presence of H. bulbosum DNA located distally on two barley chromosomes and we mapped the introgression to barley chromosome 4HL using RFLP analysis. Furthermore, by means of an allelism test we found that the transferred gene for pubescence is allelic or closely linked to a gene for pubescence (Hs) located on barley chromosome 4HL. The presence of a single H. bulbosum chromosome in the monosomic substitution plant was confirmed by GISH. A distal introgression of H. bulbosum DNA was also observed on one barley chromosome, which was located on chromosome 3HL by RFLP analysis.     

Recent progress in barley improvement using wild species of Hordeum

In this review we describe recent progress in barley (Hordeum vulgare) improvement through hybridisation with its wild relatives. We have focused on one species in the secondary genepool of cultivated barley, namely H. bulbosum. This wild species has desirable traits, such as disease resistance, that are worthwhile transferring to its cultivated relative. Progress has been achieved through developing partially fertile interspecific hybrids that have been selfed or backcrossed to barley. We present the results of cytogenetic and molecular analyses that have enabled us to characterise and produce agronomically useful recombinant lines obtained from the hybrids.     

The genomic organization and localization on the chromosomes of the HvRT family of DNA repetitive sequences in barley

HvRT family of repetitive DNA sequences from barley genome appears to have complex hierarchical organization. Tandem repetition of 118-bp monomers constitutes lower level of HvRT-family organization. Amplification units of the higher level consist of several contiguous 118-bp monomers. RFLP between different species and cultivars of barley resulted from the differences in the higher-order repeat structure. Individual chromosomes of barley contain specific HvRT subfamilies. This family also possesses separate domains differing in the restriction enzyme sites density. HvRT family is presented in the genomes of H. vulgare, H. leporinum, H. murinum, H. jubatum, but is absent in the genomes of H. marinum, H. geniculatum and wheat.     

RAPD-marking of genomes of representative Hordeum species

Random amplification of polymorphic DNA (RAPD) was used to analyze six species, three populations, and seven regional cultivars of barley. A unique pattern of amplified DNA products was obtained for each species of the genus Hordeum. High polymorphism of barley species was revealed. Specific fragments were found in most RAPD patterns; the fragments can be used as molecular markers of corresponding species and subspecies. Several other DNA fragments were shown to serve as molecular markers of the H genome. Specific RAPD patterns were obtained for each population and each cultivar of H. vulgare sensu lato. In total, variation between the populations and between the cultivars was substantially lower than between species. Cluster analysis (UPGMA) was used to estimate genetic distances between the Hordeum species, between the H. spontaneum populations, and between regional H. vulgare cultivars and a dendrogram was constructed.     

Use of aphid stylectomy and RT-PCR for the detection of transporter mRNAs in sieve elements

Unmodified samples of barley (Hordeum vulgare) sieve tube sap have been obtained by severing the stylets (stylectomy) of feeding aphids and collecting the exuding liquid. Primers were designed to direct the amplification of a series of specific cDNAs encoding barley proteins selected because of their significance in sieve tube function. mRNA encoding the H(+)/sucrose co-transporter SUT1, a putative aquaporin and the H(+)/ATPase PPA1 were detected in sieve tube sap. These mRNA species appear to be present at very low concentrations. mRNA encoding the potassium transporter HAK1 could not be detected. The results strongly suggest that some mRNA species are imported into sieve elements, which are enucleate, from neighbouring companion cells.     

Chromosome 5H of <Emphasis Type="Italic">Hordeum</Emphasis> species involved in reduction in grain hardness in wheat genetic background

Grain hardness is an important factor affecting end-use quality in wheat. Mutations of the puroindoline genes, which are located on chromosome 5DS, control a majority of grain texture variations. Hordoindoline genes, which are the puroindoline gene homologs in barley, are located on chromosome 5HS and are also responsible for grain texture variation. In this study, we used three types of wheat–barley species (Hordeum vulgare, H. vulgare ssp. spontaneum, and H. chilense) chromosome addition lines and studied the effect of chromosome 5H of these species on wheat grain characteristics. The 5H chromosome addition lines showed significantly lower grain hardness and higher grain weight than the corresponding wheat parents. The effect of enhancing grain softness was largest in the wheat–H. chilense line regardless of having an increase in grain weight similar to those in the wheat–H. vulgare and wheat–H. spontaneum lines. Our results indicated that chromosome 5H of the Hordeum species plays a role in enhancing grain softness and increasing grain weight in the wheat genetic background, and the extent of effect on grain hardness depends on the type of Hordeum species. Protein analysis of hordoindolines indicated that profiles of 2D-electrophoresis of hordoindolines were different among Hordeum species and hordoindolines in the addition lines appeared to be most abundant in wheat–H. chilense line. The differences in enhancing grain softness among the Hordeum species might be attributed to the quantity of hordoindolines expressed in the 5H chromosome addition lines. These results suggested that the barley hordoindolines located on chromosome 5HS play a role in reducing grain hardness in the wheat genetic background.