Wheat Sequences Research Articles

Wheat (Triticum aestivum) productivity under different tillage practices and legume options in rice (Oryza sativa) and wheat cropping sequence

Screening of suitable tillage practices and legumes between rice (Oryza sativa L.) and wheat (Triticum aestivum L.) is the need of hour to maintain the wheat productivity, which is hampering in rice wheat sequence. Considering this, experiments were conducted during 2006-07 to 2009-10, at Norman E Borlogue Crop Research Centre of G B Pant University of Agriculture & Technology, Pantnagar. The experimental plots were silty clay loam, medium in organic carbon, available phosphorus and low in nitrogen, however the pH was slightly alkaline. Pooled analyses were done for four years considering the years as main plot of split plot design. However, sub plot contained three tillage practices, i.e zero tillage (ZT), raised bed system (RBS) and conventional tillage (CT). Three legumes namely greengram (Vigna radiata L.), cowpea [Vigna unguiculata (L.) Walp.] and vegetable pea (Pisum sativum var hortense) were kept in sub sub plots. The findings revealed that, over the years, wheat yield markedly influenced by maximum temperature as compared to other weather attributes. Simultaneously, tillage and legume interaction was non-significant on wheat yield. Mean effect of the treatments vindicated that metric parameters, viz plant height, dry matter accumulation, biological yield, grain yield, straw yield, grain straw ratio and harvest index were significantly influenced by years, tillage practices and legume inclusion. Year 2007-08 was the good year for wheat performance. Among the tillage practices conventional tillage retained maximum metric traits than the other tillage practices, however in case of legume inclusion; cowpea gave maximum metric traits. Significant effects of treatments were also noticed on NPK uptake, available N in soil, net returns and benefit cost ratio and system productivity. Maximum values of these parameters were noticed during 2007-08. Maximum available N, NPK uptake were observed in zero tillage. Greengram containing plots had maximum available N but vegetable containing plots had higher nutrient content and uptake. Net return, benefit cost ratio were maximum under zero tillage and vegetable pea containing plots. Conventional tillage and vegetable pea contained plots gave maximum system productivity. Therefore, conventional tillage and cowpea inclusion is good for wheat yield but for economics point of view zero tillage practices and inclusion of vegetable pea is better.

The reducing end sequence of wheat endosperm cell wall arabinoxylans

Walls from wheat (Triticum aestivum L.) endosperm are composed primarily of hetero-(arabino)xylans (AXs) (70%) and (1→3)(1→4)-β-d-glucans (20%) with minor amounts of cellulose and heteromannans (2% each). To understand the differential solubility properties of the AXs, as well as aspects of their biosynthesis, we are sequencing the xylan backbone and examining the reducing end (RE) sequence(s) of wheat (monocot) AXs. A previous study of grass AXs (switchgrass, rice, Brachypodium, Miscanthus and foxtail millet) concluded that grasses lacked the comparable RE glycosyl sequence (4-β-d-Xylp-(1→4)-β-d-Xylp-(1→3)-α-l-Rhap-(1→2)-α-d-GalpA-(1→4)-d-Xylp) found in dicots and gymnosperms but the actual RE sequence was not determined.Here we report the isolation and structural characterisation of the RE oligosaccharide sequence(s) of wheat endosperm cell wall AXs. Walls were isolated as an alcohol-insoluble residue (AIR) and sequentially extracted with hot water (W-sol Fr) and 1M KOH containing 1% NaBH4 (KOH-sol Fr). Detailed structural analysis of the RE oligosaccharides was performed using a combination of methylation analysis, MALDI-TOF-MS, ESI-QTOF-MS, ESI-MSn and enzymic analysis. Analysis of RE oligosaccharides, both 2AB labelled (from W-sol Fr) and glycosyl-alditol (from KOH-sol Fr), revealed that the RE glycosyl sequence of wheat endosperm AX comprises a linear (1→4)-β-d-Xylp backbone which may be mono-substituted with either an α-l-Araf residue at the reducing end β-d-Xylp residue and/or penultimate RE β-d-Xyl residue; β-d-Xylp-(1→4)-[α-l-Araf-(1→3)](+/−)-β-d-Xylp-(1→4)-[α-l-Araf-(1→3)](+/−)-β-d-Xylp and/or an α-d-GlcpA residue at the reducing end β-d-Xylp residue; β-d-Xylp-(1→4)-[α-l-Araf-(1→3)](+/−)-β-d-Xylp-(1→4)-[α-d-GlcAp-(1→2)]-β-d-Xylp. Thus, wheat endosperm AX backbones lacks the RE sequence found in dicot and gymnosperm xylans; a finding consistent with previous reports from other grass species.

Gene Identification Programs in Bread Wheat: A Comparison Study

Seven ab initio web-based gene prediction programs (i.e., AUGUSTUS, BGF, Fgenesh, Fgenesh+, GeneID, Genemark.hmm, and HMMgene) were assessed to compare their prediction accuracy using protein-coding sequences of bread wheat. At both nucleotide and exon levels, Fgenesh+ was deduced as the superior program and BGF followed by Fgenesh were resided in the next positions, respectively. Conversely, at gene level, Fgenesh with the value of predicting more than 75% of all the genes precisely, concluded as the best ones. It was also found out that programs such as Fgenesh+, BGF, and Fgenesh, because of harboring the highest percentage of correct predictive exons appear to be much more applicable in achieving more trustworthy results, while using both GeneID and HMMgene the percentage of false negatives would be expected to enhance. Regarding initial exon, overall, the frequency of accurate recognition of 3′ boundary was significantly higher than that of 5′ and the reverse was true if terminal exon is taken into account. Lastly, HMMgene and Genemark.hmm, overall, presented independent tendency against GC content, while the others appear to be slightly more sensitive if GC-poor sequences are employed. Our results, overall, exhibited that to make adequate opportunity in acquiring remarkable results, gene finders still need additional improvements.

Next-Generation Survey Sequencing and the Molecular Organization of Wheat Chromosome 6B

Common wheat (Triticum aestivum L.) is one of the most important cereals in the world. To improve wheat quality and productivity, the genomic sequence of wheat must be determined. The large genome size (∼17 Gb/1 C) and the hexaploid status of wheat have hampered the genome sequencing of wheat. However, flow sorting of individual chromosomes has allowed us to purify and separately shotgun-sequence a pair of telocentric chromosomes. Here, we describe a result from the survey sequencing of wheat chromosome 6B (914 Mb/1 C) using massively parallel 454 pyrosequencing. From the 4.94 and 5.51 Gb shotgun sequence data from the two chromosome arms of 6BS and 6BL, 235 and 273 Mb sequences were assembled to cover ∼55.6 and 54.9% of the total genomic regions, respectively. Repetitive sequences composed 77 and 86% of the assembled sequences on 6BS and 6BL, respectively. Within the assembled sequences, we predicted a total of 4798 non-repetitive gene loci with the evidence of expression from the wheat transcriptome data. The numbers and chromosomal distribution patterns of the genes for tRNAs and microRNAs in wheat 6B were investigated, and the results suggested a significant involvement of DNA transposon diffusion in the evolution of these non-protein-coding RNA genes. A comparative analysis of the genomic sequences of wheat 6B and monocot plants clearly indicated the evolutionary conservation of gene contents.

Draft genome of the wheat A-genome progenitor Triticum urartu

Bread wheat (Triticum aestivum, AABBDD) is one of the most widely cultivated and consumed food crops in the world. However, the complex polyploid nature of its genome makes genetic and functional analyses extremely challenging. The A genome, as a basic genome of bread wheat and other polyploid wheats, for example, T. turgidum (AABB), T. timopheevii (AAGG) and T. zhukovskyi (AAGGA(m)A(m)), is central to wheat evolution, domestication and genetic improvement. The progenitor species of the A genome is the diploid wild einkorn wheat T. urartu, which resembles cultivated wheat more extensively than do Aegilops speltoides (the ancestor of the B genome) and Ae. tauschii (the donor of the D genome), especially in the morphology and development of spike and seed. Here we present the generation, assembly and analysis of a whole-genome shotgun draft sequence of the T. urartu genome. We identified protein-coding gene models, performed genome structure analyses and assessed its utility for analysing agronomically important genes and for developing molecular markers. Our T. urartu genome assembly provides a diploid reference for analysis of polyploid wheat genomes and is a valuable resource for the genetic improvement of wheat.

Comparative analysis of protein-protein interactions in the defense response of rice and wheat

BackgroundDespite the importance of wheat as a major staple crop and the negative impact of diseases on its production worldwide, the genetic mechanisms and gene interactions involved in the resistance response in wheat are still poorly understood. The complete sequence of the rice genome has provided an extremely useful parallel road map for genetic and genomics studies in wheat. The recent construction of a defense response interactome in rice has the potential to further enhance the translation of advances in rice to wheat and other grasses. The objective of this study was to determine the degree of conservation in the protein-protein interactions in the rice and wheat defense response interactomes. As entry points we selected proteins that serve as key regulators of the rice defense response: the RAR1/SGT1/HSP90 protein complex, NPR1, XA21, and XB12 (XA21 interacting protein 12).ResultsUsing available wheat sequence databases and phylogenetic analyses we identified and cloned the wheat orthologs of these four rice proteins, including recently duplicated paralogs, and their known direct interactors and tested 86 binary protein interactions using yeast-two-hybrid (Y2H) assays. All interactions between wheat proteins were further tested using in planta bimolecular fluorescence complementation (BiFC). Eighty three percent of the known rice interactions were confirmed when wheat proteins were tested with rice interactors and 76% were confirmed using wheat protein pairs. All interactions in the RAR1/SGT1/ HSP90, NPR1 and XB12 nodes were confirmed for the identified orthologous wheat proteins, whereas only forty four percent of the interactions were confirmed in the interactome node centered on XA21. We hypothesize that this reduction may be associated with a different sub-functionalization history of the multiple duplications that occurred in this gene family after the divergence of the wheat and rice lineages.ConclusionsThe observed high conservation of interactions between proteins that serve as key regulators of the rice defense response suggests that the existing rice interactome can be used to predict interactions in wheat. Such predictions are less reliable for nodes that have undergone a different history of duplications and sub-functionalization in the two lineages.

WRKY Transcription Factors in Wheat and Their Induction by Biotic and Abiotic Stress

WRKY proteins constitute a large family of transcription factors in higher plants, and are involved in the regulation of development, senescence and stress resistance. However, little is known with respect to the constitution and function of WRKY genes in bread wheat (Triticum aestivum). We identified a set of 92 wheat WRKY genes from publicly available sequence databases, and constructed a phylogeny map based on the conserved WRKY domain peptide sequences. The wheat sequences were classified into the standard set of WRKY groups and sub-groups established from plant model genomes. Our results show the WRKY orthologues of wheat and rice are more similar compared to their Arabidopsis counterparts. Therefore, we argue some expansion of the WRKY family occurred following the divergence of these three species. The induction of sequence variation in a subset of 18 TaWRKY genes via somatic hybridization was studied by comparing the sequences from cv. JN177 and its derivative cv. SR3. We also analyzed the responses of these 18 genes to imposed salinity, PEG, ABA and SA using quantitative real time PCR. Nine of the 18 TaWRKY genes were up-regulated by exogenous SA. Six genes were up-regulated by salinity or PEG treatment, and their expression was also induced by exogenous ABA. Interestingly, induction by salinity or PEG was impaired when ABA biosynthesis was inhibited. The indication was that most of the TaWRKY genes involved in the abiotic stress response acted in an ABA-dependent manner.

Genomic and chromosomal distribution patterns of various repeated DNA sequences in wheat revealed by a fluorescence in situ hybridization procedure

Wheat (Triticum aestivum L.) is an allohexaploid, in which each of the three genomes has a high 1C content. This indicates the presence of multiple tandemly repeated sequences, which should be detectable using in situ hybridization. Some repeats have already been described, but others remain to be recognized. To discover others, 2000 plasmid wheat clones were examined for signal presence after fluorescence in situ hybridization and microscopic signal observation. Among them, 47 clones produced strong discrete signals on wheat chromosomes. Two of the newly identified clones (pTa-535 and pTa-713) were determined to have especially valuable sequences for chromosome identification. In combination with pTa-86 (the pSc119 homologous sequence), these probes enable unambiguous discrimination of all wheat chromosomes including orientation. Four newly identified sequences (pTa-465, pTa-k566, pTa-s120, and pTa-s126) were useful in that they produced discrete signals on various wheat chromosome arms. Two other clones (pTa-k288 and pTa-k229) produced GISH-like (genomic in situ hybridization) signals because they allowed the A, B, and D genomes to be distinguished simultaneously. In addition, centromere, centromere-related, and ribosomal DNA clones were identified. Also described are improvements on slide preparation and reprobing procedures. To enhance discrete signal detection, a new direct fluorescent-labeling procedure, namely the VentR (exo-) terminal extension method, was employed.

Wheat centromeric retrotransposons: the new ones take a major role in centromeric structure

The physical map of the hexaploid wheat chromosome3B was screened using centromeric DNA probes. A 1.1-Mb region showing the highest number of positive bacterial artificial chromosome (BAC) clones was fully sequenced and annotated, revealing that 96% of the DNA consisted of transposable elements, mainly long terminal repeat (LTR) retrotransposons (88%). Estimation of the insertion time of the transposable elements revealed that CRW (also called Cereba) and Quinta are the youngest elements at the centromeres of common wheat (Triticumspp.) and its diploid ancestors, with Quinta being younger than CRW in both diploid and hexaploid wheats. Chromatin immunoprecipitation experiments revealed that both CRW and Quinta families are targeted by the centromere-specific histone H3 variant CENH3. Immuno colocalization of retroelements and CENH3 antibody indicated that a higher proportion of Quinta than CRWs was associated with CENH3, although CRWs were more abundant. Long arrays of satellite repeats were also identified in the wheat centromere regions, but they lost the ability to bind with CENH3. In addition to transposons, two functional genes and one pseudogene were identified. The gene density in the centromere appeared to be between three and four times lower than the average gene density of chromosome3B. Comparisons with related grasses also indicated a loss of microcollinearity in this region. Finally, comparison of centromeric sequences of Aegilops tauschii (DD), Triticum boeoticum (AA) and hexaploid wheat revealed that the centromeres in both the polyploids and diploids are still undergoing dynamic changes, and that the new CRWs and Quintas may have undertaken the core role in kinetochore formation.

Genetic characterization and mapping of the Rht-1 homoeologs and flanking sequences in wheat

The introgression of Reduced height (Rht)-B1b and Rht-D1b into bread wheat (Triticum aestivum) varieties beginning in the 1960s led to improved lodging resistance and yield, providing a major contribution to the 'green revolution'. Although wheat Rht-1 and surrounding sequence is available, the genetic composition of this region has not been examined in a homoeologous series. To determine this, three Rht-1-containing bacterial artificial chromosome (BAC) sequences derived from the A, B, and D genomes of the bread wheat variety Chinese Spring (CS) were fully assembled and analyzed. This revealed that Rht-1 and two upstream genes were highly conserved among the homoeologs. In contrast, transposable elements (TEs) were not conserved among homoeologs with the exception of intronic miniature inverted-repeat TEs (MITEs). In relation to the Triticum urartu ancestral line, CS-A genic sequences were highly conserved and several colinear TEs were present. Comparative analysis of the CS wheat BAC sequences with assembled Poaceae genomes showed gene synteny and amino acid sequences were well preserved. Further 5' and 3' of the wheat BAC sequences, a high degree of gene colinearity is present among the assembled Poaceae genomes. In the 20kb of sequence flanking Rht-1, five conserved non-coding sequences (CNSs) were present among the CS wheat homoeologs and among all the Poaceae members examined. Rht-A1 was mapped to the long arm of chromosome 4 and three closely flanking genetic markers were identified. The tools developed herein will enable detailed studies of Rht-1 and linked genes that affect abiotic and biotic stress response in wheat.

Integrating cereal genomics to support innovation in the Triticeae.

The genomic resources of small grain cereals that include some of the most important crop species such as wheat, barley, and rye are attaining a level of completion that now is contributing to new structural and functional studies as well as refining molecular marker development and mapping strategies for increasing the efficiency of breeding processes. The integration of new efforts to obtain reference sequences in bread wheat and barley, in particular, is accelerating the acquisition and interpretation of genome-level analyses in both of these major crops.

Influences of crop sequence, rainfall and irrigation, on relationships between Gaeumannomyces graminis var. tritici and take-all in New Zealand wheat fields

In a study, over three growing seasons, of Gaeumannomyces graminis var. tritici (Ggt) and take-all disease in commercial wheat fields (that were in their first, second, third or fourth year of consecutive wheat) in New Zealand, Ggt concentrations in soil (the amount of Ggt DNA measured using quantitative PCR) and take-all incidence and take-all index (TAI) were least at the start of a wheat sequence and stabilized in third and fourth consecutive wheat crops. Median Ggt concentrations in soil increased 10-fold during a first wheat crop, reaching 78 pg Ggt DNA/g soil after harvest. Growing season rainfall and position in a continuous wheat crop sequence were most closely associated with TAI and post-harvest soil Ggt concentrations, but other factors were also associated with inoculum and disease. Inoculum concentrations, disease incidence and TAI were greater where the frequency of crops susceptible to Ggt (wheat, barley or triticale) was greatest in the crop rotations and where crops were irrigated. Irrigation in particular was associated with high post-harvest soil Ggt concentrations in the driest of the three growing seasons assessed, when environmental conditions were least favourable for disease development. There was evidence of take-all decline following two wheat crops where environmental conditions had enabled severe epidemics to occur. Our study showed that frequency of cereal crops in the rotation, length of breaks between host crops, irrigation and growing season rainfall interact over the course of a cropping sequence to influence TAI when wheat is sown.

Root-lesion nematode (Pratylenchus thornei) reduces nutrient response, biomass and yield of wheat in sorghum–fallow–wheat cropping systems in a subtropical environment

Wheat in winter and grain sorghum in summer are the most important crops of the subtropical grain region of eastern Australia, where they are often grown in sequences of 1–4 years of each crop species separated by a long fallow period of 11–14 months. Growers have observed that the second wheat crop in sequence can appear nutrient deficient with poor growth and grain yield compared with the first or subsequent crops. To investigate this problem, wheat was grown in various field experiments as the first–fourth wheat in sequence after long fallow from sorghum sequences of various lengths (one–four years) to test for responses in biomass production and grain yield to fertilisers, biocidal fumigants and nematicides. Populations of root-lesion nematodes (Pratylenchus thornei) were greatest in the soil before the second wheat crop was sown than before the first or third wheat crops. Greatest responses in wheat growth were obtained to the nematicide aldicarb in various wheat sequence positions up to the fourth (up to 137% increased yield of second wheat). Aldicarb best protected the roots from P. thornei allowing increased nutrient response. The fumigants chloropicrin and dazomet caused substantial changes in soil microbial populations and available nutrients, but the systemic nematicides fenamiphos and aldicarb did not. Responses in grain yield were also obtained to N fertiliser and less frequently to P and Zn (41% increase of second wheat to NPZn fertiliser), with best overall responses to a combination of aldicarb and fertiliser. A range of tolerance to P. thornei, as judged by both grain yield from nil treatment and response to aldicarb and/or fertiliser, was identified among wheat cultivars. In comparison, one barley cultivar yielded maximally without treatment (up to 3.7 and 1.3 times the yield of the most intolerant and the most tolerant wheat cultivars, respectively). Integrated management using crop rotation with sorghum to reduce P. thornei populations combined with growing tolerant wheat and barley cultivars supplied with adequate fertiliser are practical measures to reduce the impact of P. thornei.

Construction and analysis of suppression subtractive hybridization library for Leymus secalinus exposed to drought stress

The response of Leymus secalinus under drought stress was studied using the method of suppression subtractive hybridization (SSH). The subtracted complementary deoxyribonucleic acid (cDNA) library containing 539 clones for leaves of L. secalinus exposed to drought stress was screened by dot-blot hybridization. Among these clones, 16 ESTs were identified as drought stress induced genes and 13 genes were identified from them. Verification by quantitative real time polymerase chain reaction (qPCR), confirmed that ten genes were up-regulated genes under drought stress and three genes had no obvious fold changes under drought stress condition. The BLAST results show that these sequences are highly homologous to the gene sequence of wheat and barley under drought stress. The genes related to drought stress play an important roles in controlling plant osmotic pressure, heat adjustment, photosynthesis and hydrolysis. This research laid a solid foundation for further study of molecular mechanism of drought resistance in L. secalinus. Key words: Leymus secalinus, drought stress induced genes, suppression subtractive hybridization (SSH), complementary deoxyribonucleic acid (cDNA), quantitative real time polymerase chain reaction (qPCR).

Site-specific wild oat (Avena fatua L.) management

Beckie, H. J. and Shirriff, S. 2012. Site-specific wild oat ( Avena fatua L.) management. Can. J. Plant Sci. 92: 923–931. Variation in soil properties, such as soil moisture, across a hummocky landscape may influence wild oat emergence and growth. To evaluate wild oat emergence, growth, and management according to landscape position, a study was conducted from 2006 to 2010 in a hummocky field in the semiarid Moist Mixed Grassland ecoregion of Saskatchewan. The hypothesis tested was that wild oat emergence and growth would be greater in lower than upper slope positions under normal or dry early growing season conditions. Three herbicide treatments were imposed on the same plots each year of a 2-yr canola (Brassica napus L.) – wheat (Triticum aestivum L.) sequence: (1) nontreated (weedy) control; (2) herbicide application to upper and lower slope positions (i.e., full or blanket application); and (3) herbicide application to lower slope position only. Slope position affected crop and weed densities before in-crop herbicide application in years with dry spring growing conditions. Site-specific wild oat herbicide application in hummocky fields in semiarid regions may be justified based on results of wild oat control averaged across slope position. In year 2 of the crop sequence (wheat), overall (i.e., lower and upper slope) wild oat control based on density, biomass, and dockage (i.e., seed return) was similar between site-specific and full herbicide treatment in 2 of 3 yr. Because economic thresholds have not been widely adopted by growers in managing wild oat, site-specific treatment in years when conditions warrant may be an appropriate compromise between no application and blanket herbicide application.

Pulse Crops Improve Energy Intensity and Productivity of Cereal Production in Montana, USA

Energy consumption, intensity, and productivity are indicators of agricultural sustainability in the face of fossil energy scarcity and price volatility. In this study, budgets of energy embodied in crop production inputs were compiled for 14 paired pulse (annual grain legume)-wheat and wheat-wheat crop sequences using data collected from Montana farmers. We report two energy performance metrics: net energy yield (NEY), the energy content of harvested crop minus energy required to produce it, and energy intensity (EI), the ratio of input energy to mass of crop harvested. Nitrogen fertilizer accounted for 58% of the energy used in wheat production, and its absence largely accounted for the 53% reduction in energy inputs to pulses relative to wheat. EI of pulses was lower than for wheat, and pulse crops also resulted in reduced EI and increased NEY for the subsequent wheat crop compared to wheat grown following wheat. The largest component of the improved energy performance of wheat following pulses was increased yield rather than decreased inputs.

The WRKY transcription factor family in Brachypodium distachyon

BackgroundA complete assembled genome sequence of wheat is not yet available. Therefore, model plant systems for wheat are very valuable. Brachypodium distachyon (Brachypodium) is such a system. The WRKY family of transcription factors is one of the most important families of plant transcriptional regulators with members regulating important agronomic traits. Studies of WRKY transcription factors in Brachypodium and wheat therefore promise to lead to new strategies for wheat improvement.ResultsWe have identified and manually curated the WRKY transcription factor family from Brachypodium using a pipeline designed to identify all potential WRKY genes. 86 WRKY transcription factors were found, a total higher than all other current databases. We therefore propose that our numbering system (BdWRKY1-BdWRKY86) becomes the standard nomenclature. In the JGI v1.0 assembly of Brachypodium with the MIPS/JGI v1.0 annotation, nine of the transcription factors have no gene model and eleven gene models are probably incorrectly predicted. In total, twenty WRKY transcription factors (23.3%) do not appear to have accurate gene models. To facilitate use of our data, we have produced The Database of Brachypodium distachyon WRKY Transcription Factors. Each WRKY transcription factor has a gene page that includes predicted protein domains from MEME analyses. These conserved protein domains reflect possible input and output domains in signaling. The database also contains a BLAST search function where a large dataset of WRKY transcription factors, published genes, and an extensive set of wheat ESTs can be searched. We also produced a phylogram containing the WRKY transcription factor families from Brachypodium, rice, Arabidopsis, soybean, and Physcomitrella patens, together with published WRKY transcription factors from wheat. This phylogenetic tree provides evidence for orthologues, co-orthologues, and paralogues of Brachypodium WRKY transcription factors.ConclusionsThe description of the WRKY transcription factor family in Brachypodium that we report here provides a framework for functional genomics studies in an important model system. Our database is a resource for both Brachypodium and wheat studies and ultimately projects aimed at improving wheat through manipulation of WRKY transcription factors.

Isolation and characterisation of a banana CYCD2;1 gene and its over-expression enhances root growth

Stimulation of the cell cycle by over-expression of Arabidopsis CyclinD2;1 enhanced growth in tobacco and rice, but not Arabidopsis. To broaden the range of species and to improve an important crop species for developing countries, a CyclinD2;1 homologue was isolated from an East African highland banana cultivar ‘Nakasabira’ ( Musa spp., AAA group) and designated Musac;CYCD2;1 (Genebank accession no HQ839770). Musac;CYCD2;1 amino-acid sequence exhibited 40, 54, 57 and 57% identity with CYCD2;1 sequences of Arabidopsis, rice, maize and wheat, respectively. Over-expression of Musac;CYCD2;1 driven by the CaMV 35S promoter in cultivar ‘Sukali ndizi’ (AAB) resulted in transformed plants with no significant increase in total banana CYCD2;1 transcripts in the shoot, but up to 66-fold increase of total banana CYCD2;1 transcripts in roots. Correspondingly, transformed plants had no change in above-ground growth but produced longer main roots and more lateral roots under in vitro conditions. A deeper root system was observed in one transgenic line relative to non-transformed plants when grown in soil. Results demonstrate that a banana homologue of Arabidopsis CYCD2;1 can also enhance growth in a homologous background and the potential of enhancing banana root growth. Key words: CyclinD2;1 gene, Musa spp., banana transformation, CyclinD2;1 over-expression, root growth.

Next generation sequencing of DNA in 3300-year-old charred cereal grains

We investigated whether ‘next generation’ methods can be used to sequence ancient DNA molecules in charred cereal grains. We prepared a DNA extract from a mixed sample of barley, einkorn, emmer and broomcorn millet, taken from a 3300-year-old assemblage of charred cereal grains from Assiros Toumba, Greece. Using the SOLiD 5500 system, we obtained 21,112,844 unique sequence reads. Of these, 178,779 had a 75% or greater nucleotide sequence similarity with one or more entries in the full nucleotide sequence database; 496 of these matches were to previously reported sequences from barley, einkorn, emmer, broomcorn millet or related species such as hexaploid wheat. The unique reads were also compared with a database comprising only wheat sequences. This analysis identified 1658 charred grain sequences that had 90% or greater similarity with segments of the wheat genome. The presence of barley, wheat and millet sequences in the next generation dataset confirms the presence of ancient DNA in this charred grain assemblage. Enrichment of extracts by hybridization capture or equivalent methods is likely to enable sequences to be obtained for entire genes and other genomic regions of interest.

Loss of Parental Coding Sequences in an Early Generation of Wheat-Rye Allopolyploid

During wheat-rye allopolyploidization, the characteristics of the sequences in the bands that appear in both parents and progeny are still unclear. In this study, two different combinations of wheat (Triticum aestivum L.) and rye (Secale cereale L.), including F1 hybrids and the first and second allopolyploid generations, were analyzed by PCR and sequencing using 60 wheat expressed sequence tag (EST)–derived single-sequence repeat markers and EST-derived sequence-tagged site markers. Thirty markers produced the same bands from parental plants, F1 plants, and amphiploids. Seven of the 30 markers amplified identical sequences from wheat and rye parents. Most of these sequences have high similarity between the two parental plants. The variation patterns of sequences in the bands produced by the seven markers were observed. In the F1 hybrids and amphiploids, loss of parental sequences was observed and the frequency of losing rye sequences was higher than that of losing wheat sequences. In addition, a few sequences in these bands exhibited significant differences, indicating that parental sequences changed drastically during allopolyploidization. Therefore, the fact that the parents and progeny contained the same bands should not be regarded as conservation. The results in this study add to the investigations dealing with variation patterns of coding sequences during wheat-rye allopolyploidization.