Molecular cloning and characterization of six novel HMW‐GS genes from Aegilops speltoides and Aegilops kotschyi

Six novel high molecular weight glutenin subunits (HMW‐GS) from Aegilops speltoides (SS, 2n = 2x = 14) and Aegilops kotschyi (UUSS, 2n = 4x = 28) were identified by SDS‐PAGE and designated as ASy15*, AKx1*, AKx3*, AKx2.3, AKy20* and AKy8*, respectively. Their complete open reading frames (ORFs) were cloned and sequenced by allele‐specific PCR (AS‐PCR). Sequence comparison demonstrated that these novel genes displayed high single‐nucleotide polymorphisms (SNP) and InDel variations. In particular, AKy8* had an extra cysteine residue at position 140 in the central repetitive domain, while AKx2.3 had an unusually long repetitive domain (816 aa), which might have positive effects on gluten quality. Phylogenetic analysis showed that AKx1* and AKx3* belonged to the 1S genome, AKx2.3 to the 1U genome, and AKy20* and AKy8* most likely to the 1S genome. The divergence between the 6 HMW‐GS genes from the two Aegilops species and those from Triticum species occurred 4.77–23.54 MYA. The authenticity of these isolated endogenous HMW‐GS genes was confirmed through heterologous expression in Escherichia coli and Western blotting.     

Molecular advancements in male sterility systems of Capsicum: A review

In recent years, plant molecular research on genetic mapping, gene tagging and cloning, and marker-assisted selection (MAS) have gained importance in crop improvement programmes. In Capsicum, several inter- and intra-specific genetic maps with wide distribution of markers covering the whole genome have been developed. Recently, whole genome of the hot pepper C. annuum, its wild progenitor C. annuum var. glabriusculum and C. baccatum has been sequenced. The Capsicum genome size has been estimated to be approx. 4× (3.48 Gb) the genome size of cultivated tomato (Solanum lycopersicum L.) (900 Mb). Breeders’ access to the pepper genomic information would facilitate the choice of markers from different linkage groups, thus paving the way for gene cloning and its introgression into the elite breeding lines through MAS. Till date, approx. 20 independently inherited nuclear male sterility (NMS) genes have been reported. Linked markers have been identified for ms1, ms3, ms8, ms10, ms_k, msc-1 and an undesignated gene. However, markers tightly linked to ms8 and ms10 are still lacking. Except ms1, ms3, ms8 and ms10, the map position of other NMS genes is not known. In cytoplasmic male sterility (CMS), markers for the mitochondrial gene atp6 have been developed and the gene cloned. Number of markers some very tightly linked to the restorer-of-fertility (Rf) gene have been identified. However, the actual map position of the Rf locus is still not determined. Another CMS-associated nuclear gene “pr” responsible for restoring partial fertility has been identified and tagged. In this review, we have compiled up-to-date information about the marker technology relating to the NMS and the CMS-associated genes in Capsicum. This information can be useful when screening Capsicum germplasm, developing NMS lines through MAS, improving efficiency of the NMS system, transferring rf gene for maintainer line breeding and Rf genes for restorer line breeding in CMS and assessing genetic purity of the hybrid seed.     

Ancient Polyploidy and Modem Crop Plants

The growing set of fully-sequenced angiosperm genomes highlight the role of polyploidy in angiosperm evolution, and suggest that even the high level of importance we had already attributed to this mechanism was inadequate. Gene loss following whole-genome duplication events has contributed much to the incongruities that we observe in comparative genetic maps, and presumably has also contributed to functional divergence of taxa. Ancient genome duplication has several consequences for levels and patterns of variation in modern plant populations, in particular contributing to the relatively conservative evolution of duplicated genes even at distant locations from one another in the genome. Comparison of gene retention/loss following independent whole-genome duplication events suggests patterns that may be relevant to the survival and productivity of newly-formed polyploids, with numerous implications for crop improvement.     

Complete mitochondrial genome sequence of Brassica oxyrrhina and comparative analysis of the region containing orf108, a male sterility gene for mustard (Brassica juncea), among B. oxyrrhina,Diplotaxis erucoides and Sinapis species

To date, several cytoplasms of wild species have been introduced to Brassica juncea, for inducing cytoplasmic male sterility (CMS). One of the causal genes of CMS is orf108, which is widely distributed in Brassicaceae including Brassica oxyrrhina. To further understand the origin of orf108, we assembled the complete mitochondrial genome sequence of B. oxyrrhina. We also determined the DNA sequences upstream of atp1 including orf108 for D. erucoides and five Sinapis species. The orf108 was definitively placed in the mitochondrial genome of B. oxyrrhina consisting of 247,936 bp. In S. alba, previously reported to possess orf108, the gene was not present, whereas Sinapis arvensis and Sinapis turgida contained orf108. The orf108 sequence in the two Sinapis species showed a novel nucleotide substitution compared to D. erucoides. Northern hybridization suggested, furthermore, that orf108 mRNA was processed in the two species similarly to that in B. oxyrrhina. The results clarified the interspecific differentiation of orf108‐apt1 region in Sinapis.     

Genetic constitution and diversity in four narrow endemic redwoods from the family Cupressaceae

The genetic constitution and diversity of four relictual redwoods are discussed in this review. These include monotypic genera of the family Cupressaceae: coast redwood (Sequoia sempervirens), giant sequoia (Sequoiadendron giganteum), dawn redwood (Metasequoia glyptostroboides), and alerce (Fitzroya cupressoides). All four species are narrow endemics, share a number of common phenotypic traits, including red wood, and are threatened species. Fossil history suggests that the ancestors of redwoods probably originated during the Cretaceous and Tertiary periods and flourished thereafter for millions of years. Towards the end of the Tertiary period began their decline and struggle for existence that continued during the subsequent geologic upheavals and climate changes, until the survival of the present-day redwoods in the current restricted locations in the world (USA, China, and South America). Although two species, Sequoiadendron and Metasequoia, are diploids (2n = 22), and the other two are polyploids: Fitzroya a tetraploid (2n = 4x = 44), and Sequoia a hexaploid (2n = 6x = 66); they all share the same basic chromosome number x = 11. The genome size in the hexaploid Sequoia is one of the largest (31,500 MB) in the conifers, while the genome sizes of diploid Metasequoia and Sequoiadendron are about one-third (~10,000 MB) of Sequoia. Genetic diversity in the redwoods is lower than most other gymnosperms, except in Sequoia, which seems to rank near the upper quarter of the coniferous forest trees. Genomic research is sparse in the redwoods, and should be pursued for a better understanding of their genome structure, function, and adaptive genetic diversity.     

Molecular structure of a novel y-type HMW glutenin subunit gene present in Triticum tauschii

An unusually small y-type high molecular weight (HMW) glutenin subunit gene from Triticum tauschii was sequenced. This gene, encoded at the Glu-D^t1 locus was designated 12.4^t and is the smallest HMW glutenin subunit gene described so far in Triticum species. Oligonucleotide primers based on published sequences of HMW glutenin genes were designed to amplify the encoding region and the central repetitive domain of the gene, which produced fragments of 1.4 and 0.85 kb, respectively. PCR products were cloned and sequenced. The derived amino acid sequence was compared with the amino acid sequences of the HMW glutenin subunits Dy12^t, from T. tauschii, and subunits Dy10 and Dy12 of T. aestivum. The amino acid sequence deduced from the nucleotide sequence demonstrated that deletions of hexapeptides and nonapeptides were responsible for the reduction in the size of this HMW glutenin subunit. The estimated molecular weight of the Dy12.4^t subunit, calculated on the basis of the deduced amino acid sequence, was 45,228 Daltons. There were also single amino acid differences in the N-, C-terminal and central repetitive domains of this gene in comparison to the three other y-type subunits encoded at the Glu-D1 locus. The Dy12.4^t subunit showed the highest similarity to the Dy12 subunit present in the hexaploid wheat Chinese Spring.     

Identification of RAPD markers linked to recessive genes conferring siliqua shatter resistance in Brassica rapa

Shattering of siliquae causes significant seed loss in canola (Brassica napus) production worldwide. There is little genetic variation for resistance to shatter in canola and, hence, the trait has been studied in B. rapa. Previous studies have shown two randomly segregating recessive genes to be responsible for shatter resistance. Three random amplified polymorphic DNA markers were identified as being linked to shatter resistance using bulked segregant analysis in a F₃B. rapa population. The population was derived from a cross between a shatter‐susceptible Canadian cultivar and a shatter‐resistant Indian line. Of the three markers, RAC‐3₉₀₀ and RX‐7₁₀₀₀ were linked to recessive sh1 and sh2 alleles, and SAC‐20₁₃₀₀ was linked to both dominant Sh1 and Sh2 alleles. The common marker for the dominant wild‐type allele for the two loci was explained to have resulted from duplication of an original locus and the associated markers through chromosome duplication and rearrangements in the process of evolution of the modern B. rapa from its progenitor that had a lower number of chromosomes. Segregation data from double heterozygous F₃ families, although limited, indicated the markers were not linked to each other and provided further evidence for the duplication hypothesis.     

Identification of a Brassica juncea-derived recessive gene conferring resistance to Leptosphaeria maculans in oilseed rape

Screening of 212 spring type Brassica napus lines carrying B genome chromosome additions and introgressions from B. nigra, B. juncea and B. carinata resulted in the identification of one line segregating for resistance to Leptosphaeria maculans (anamorph Phoma lingam) at the seedling (cotyledon) stage. This line was derived from an interspecific hybrid containing the B genome of B. juncea. Trypan blue staining of cotyledons from resistant individuals demonstrated a hypersensitive response which is delayed in plants with intermediate lesion size. Genetic analysis supported the hypothesis of a monogenic recessive inheritance of resistance. The resistance gene, termed r_jlm2, is effective in spring and winter type oilseed rape backgrounds against all tested virulent pathotypes, including two isolates which have been shown to overcome two dominant (race‐specific) B genome‐derived resistance genes in B. napus.     

S‐locus diversity and cross‐compatibility of wild Prunus avium for timber breeding

Prunus avium is primarily cultivated for its fruit, sweet cherries. However, it is also used to produce high‐quality timber. In a P. avium seed orchard, gametophytic self‐incompatibility is a restriction for free pollen flow and should be considered when establishing basic forest materials. In this study, S‐locus diversity and cross‐incompatibility of wild cherry individuals in clonal banks established for breeding for timber production were investigated. Wild cherry trees (140) with outstanding forest growth habit, collected in northern Spain, grafted and planted in two clonal banks, were genotyped at the S‐locus. The self‐incompatibility S‐locus genes, S‐RNase and SFB, were analysed by PCR. Twenty‐two S‐haplotypes, resulting in 72 different S‐genotypes, were identified. The genotypes were grouped into 33 incompatibility groups and 39 unique genotypes. This initial S‐locus analysis revealed large genetic diversity of wild cherry trees from the Spanish northern deciduous forest, and provides useful information for seed orchard design. Wild P. avium displays significantly more genetic diversity than what is detected in local cultivars, revealing a narrowing of genetic diversity during local domestication.     

Intravarietal polyploidy in the apple (Malus pumila Mill.) cultivar Hazratbali

Summary A triploid cytotype of cv. Hazratbali (Malus pumila Mill.), was found growing in orchards of diploid trees. Morphological comparisons between the two cytotypes reveal that the triploid is more vigorous, and bears more attractive fruit. The meiotic behaviour of the two cytotypes has been described. The impact of polyploidy on quality characteristics of apple at the intravarietal level is analysed. The possible mode of origin of the triploid cytotype is discussed.     

A review on transgenic approaches to accelerate breeding of woody plants

The long juvenile period of trees delays the breeding of new varieties. Flowering begins within 5–10 years in most cultivated forest trees under intensive management, but can take up to 40 years in some species and environmental conditions. To accelerate the breeding process several agrotechnical and biotechnical methods have been developed. Knowledge about genes controlling flower initiation in model plants like Arabidopsis thaliana , and identification of homologous genes in trees, have led to new possibilities for early-flower induction. Overexpression of MADS-box and other floral regulatory genes resulted in early flowering in some tree species and/or varieties. However, these methods have not yet been shown to enable the production of fertile, viable or normal gametes and progeny; developmental research towards these ends is therefore of high priority. A breeding scheme has been developed to use early flowering trees for the introduction of genes from wild species that would allow several backcrosses to occur in only a few years, and to produce at the end a non-transgenic improved variety. Research to develop practical early flowering methods could lead to several new methods for breeding and biotechnology.     

Mitotic analysis of sticky chromosomes in aluminum tolerant and susceptible wheat lines grown in soils of differing aluminum saturation

Plant resistance is currently the most effective and environmentally safe method to control plant parasitic nematodes (PPNs). Resistance genes generally act against sedentary PPNs by inducing a hypersensitive reaction that prevents the parasite installation and/or reproduction. However, the recent emergence of virulent biotypes able to overcome the plant resistance genes may constitute a severe limitation to this control strategy. In selection experiments conducted under controled environment, the genetic variation, specificity and inheritance of nematode virulence have been demonstrated. Moreover, the occurrence of gene-for-gene interactions has been shown in a few cases. Moleculars markers have been extensively used to investigate the genetic variability of PPNs, but so far, the genomic polymorphisms observed are largely independent of virulence. Such data suggest that, within a species, virulent isolates do not share a common origin, but are probably the result of independent mutational events. To understand the molecular mechanisms responsible for virulence in PPNs, several strategies have been developed, in relation with their mode of reproduction (parthenogenesis versus amphimixis). As an example, recent results obtained in our laboratory on the root-knot nematodes Meloidogyne spp. are presented. On a more general point of view, factors that may induce stable genome variability in PPNs, e.g. Transposition of mobile elements and chromosomal rearrangements (leading to polyploidy, aneuploidy, etc) will also be considered. Advances in knowledge in these areas should have important consequences for the management and durability of natural resistance genes, and for the engineering of new forms of resistance. This revised version was published online in August 2006 with corrections to the Cover Date.     

Selection for adaptation in multipurpose trees and shrubs for production and function in agroforestry systems

Summary With the greater uniformity in site conditions in cultivated fields and higher levels of inputs (soil amendments and labour) for modern agroforestry systems, selection and breeding for optimal tree adaptation to site conditions and management practices call for strategies radically different from those for plantation forestry. Furthermore, specific requirements for the most efficient growth resource sharing arrangements between the tree and its companion crop call for evaluation both above-ground and below-ground tree growth characteristics in much greater details than has been the case for forestry practice.Specific adaptations in tree-soil relations are highlighted. Tree ideotype profiles are defined for a valuable fodder species (Calliandra calothyrsus) and a valuable timber species (Grevillea robusta). Results from half-sib progeny testing experiment on C. calothyrsus in alley farming system indicate differential adaptedness to frequent cutting for fodder production among the families after 6 cutting cycles (over a period of 2 years). If this differential response to frequent cutting among genotypes is confirmed, phyllomorphs which are more tolerant to this management practice could be selected for further improvement research.Excavation of 2 trees per family within G. robusta half-sib progeny testing experiments at two locations in Kenya provided the base for studying (a) variations in root growth patterns and (b) correlations between stem and root growth patterns in the background of efficient soil resource sharing strategies and of potentials for indirect selection for root growth characteristics. Analyses based on 7 categories of root growth and 7 categories of stem growth patterns involving 94 families selected in Australia and 60 families selected from landraces in Kenya indicate (a) insignificant variation among families in root growth patterns and (b) non existent or weak correlations between stem and root growth pattenrs.     

同源多倍体化效应研究进展

同源多倍体在自然界中的分布非常广泛。为了全面了解与掌握同源多倍体的相关研究进展,明晰相关的研究重点与薄弱环节。对国内外相关研究成果进行了综合分析,重点介绍了多倍体的各种类型以及同源多倍体植物中基因表达变化的特点, 包括基因的沉默、激活和基因表达水平的变化, 探讨了基因表达变化的分子机制。另外,对基因组加倍在进化中的效应也做了阐述。研究发现,与典型的同源多倍体显著不同的是,不少二倍体化的同源多倍体基因组出现缩减的现象。有限的研究资料显示,基因组加倍后的最初几代内没有经历剧烈的基因重构,也没有经历广泛的基因组重组。生物地理学及生态学研究发现同源多倍体的产生与环境的改变有紧密的联系。与异源多倍体相比,同源多倍体的研究还没有得到应有的重视。今后应对同源多倍体基因组的大小和基因表达模式进行不同层次与系统的研究。可以预见同源多倍体在农作物品种改良及农业生产上的积极作用将非常深远。     

双胚苗水稻来源的单倍体、二倍体及其杂交F1的DNA甲基化位点分析

全基因组倍增或多倍化, 伴随着基因丢失和二倍化进程, 被认为是植物进化的重要推动力量。DNA甲基化与miRNA的表观遗传调控机制在植物生长发育及进化过程中起着重要的作用。本文采用MSAP (甲基化敏感扩增多态性)技术分析同一双胚苗水稻来源的单倍体、二倍体及其杂交F₁的基因组DNA 5'-CCGG位点胞嘧啶的甲基化及遗传特点。对部分甲基化位点进行切胶、回收、测序及功能注释, 并结合miRNA靶基因预测探讨特定甲基化位点的遗传特点及其与miRNA的相关性。16对选择性扩增引物在双亲及杂交F₁中共检测了462个DNA甲基化位点, 杂交F₁甲基化水平平均为43.20%, 与双亲相差不大(单倍体为46.75%, 二倍体为41.99%)。以TargetFinder软件分析发现其中的7个甲基化位点基因序列上存在1~4个miRNA的结合位点, 这些基因的功能注释包括逆转录转座子蛋白、ras相关蛋白、H2A/H2B/H3/H4核心组蛋白等。同时, 探讨了逆转录转座子在植物进化中的作用。研究结果为进一步阐明水稻基因组倍增过程中DNA甲基化与miRNA的关系提供了参考。     

Genetics of leaf rust resistance in three Americano landrace-derived wheat cultivars from Uruguay

A genetic analysis of the landrace‐derived wheat accessions Americano 25e, Americano 26n, and Americano 44d, from Uruguay was conducted to identify the leaf rust resistance genes present in these early wheat cultivars. The three cultivars were crossed with the leaf rust susceptible cultivar ‘Thatcher’ and approximately 80 backcross (BC1) F₂ families were derived for each cross. The BC1F₂ families and selected BC1F₄ lines were tested for seedling and adult plant leaf rust resistance with selected isolates of leaf rust, Puccinia triticina. The segregation and infection type data indicated that Americano 25e had seedling resistance genes Lr3, Lr16, an additional unidentified seedling gene, and one adult plant resistance gene that was neither Lr12 nor Lr13, and did not phenotypically resemble Lr34. Americano 26n was postulated to have genes Lr11, Lr12, Lr13, and Lr14a. Americano 44d appeared to have two possibly unique adult plant leaf rust resistance genes.     

Erucic acid heredity in Brassica juncea - some additional information

Genetic studies were undertaken to reassess erucic acid heredity in Brassica juncea. Analysis of segregation in F₂ and BC₁ generations from two zero × high erucic acid crosses indicated that higher erucic acid in B. juncea was controlled by two dominant genes with additive effects, whereas segregation in a cross involving ‘CCWF 16′, a genotype having intermediate erucic acid (25.6%), and a zero erucic acid strain, indicated monogenic dominant control for intermediate erucic acid content. The B. juncea strain ‘CCWF 16’ was developed by hybridizing high‐erucic acid B. juncea cv.‘WF‐1’ with a ‘0’ erucic B. rapa cv.‘Candle’ followed by backcrossing with ‘WF‐1’ and half‐seed selection for low erucic acid in each backcross generation. This strategy resulted in substitution of the high erucic acid allele present in the A genome of B. juncea (AABB) by the zero erucic acid allele associated with ‘A’ genome of ‘Candle’. The intermediate erucic acid content in ‘CCWF 16’ was thus attributed to a gene present in the ‘BB’ genome. Experimental data clearly suggested that the gene (E₂) associated with the A genome had a greater contribution to the total erucic acid content in B. juncea than the gene (E₁) located on the B genome. This provided experimental evidence for a previous suggestion of unequal contributions of two dominant genes (E₁= 12%, E₂= 20%) to high erucic acid content in conventional digenomic Brassica species.     

Rj (rj) genes involved in nitrogen-fixing root nodule formation in soybean

It has long been known that formation of symbiotic root nodules in soybean (Glycine max (L.) Merr.) is controlled by several host genes referred to as Rj (rj) genes, but molecular cloning of these genes has been hampered by soybean’s complicated genome structure and large genome size. Progress in molecular identification of legume genes involved in root nodule symbiosis have been mostly achieved by using two model legumes, Lotus japonicus and Medicago truncatula, that have relatively simple and small genomes and are capable of molecular transfection. However, recent development of resources for soybean molecular genetic research, such as genome sequencing, large EST databases, and high-density linkage maps, have enabled us to isolate several Rj genes. This progress has been achieved in connection with systematic utilization of the information obtained from molecular genetics of the model legumes. In this review, we summarize the current status of knowledge of host-controlled nodulation in soybean based on information from recent studies on Rj genes, and discuss the future research prospects.