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Abstract: Alternative oxidase (AOX) is the central component of the non-phosphorylating alternative respiratory pathway in plants and may be important for mitochondrial function during environmental stresses. Recently it has been proposed that Aox can be used as a functional marker for breeding stress tolerant plant varieties. This requires characterization of Aox alleles in plants with different degree of tolerance in a certain stress, affecting plant phenotype in a recognizable way. In this study we examined Aox1 gene expression levels in Medicago truncatula genotypes differing in salt stress tolerance, in order to uncover any correlation between Aox expression and tolerance to salt stress. Results demonstrated a specific induction of Aox1 gene expression in roots of the tolerant genotype that presented the lowest modulation in phenotypic and biochemical stress indices such as morphologic changes, protein level, lipid peroxidation and ROS generation. Similarly, in a previous study we reported that induction of antioxidant gene expression in the tolerant genotype contributed to the support of the antioxidant cellular machinery and stress tolerance. Correlation between expression patterns of the two groups of genes was revealed mainly in 48h treated roots. Taken together, results from both experiments suggest that M. truncatula tolerance to salt stress may in part due to an efficient control of oxidative balance thanks to (i) induction of antioxidant systems and (ii) involvement of the AOX pathway. This reinforces the conclusion that differences in antioxidant mechanisms can be essential for salt stress tolerance in M. truncatula and possibly the corresponding genes, especially Aox, could be utilized as functional marker. [Copyright &y& Elsevier]

Abstract: Molecular innovations in microbial ecology are allowing scientists to correlate microbial community characteristics to a variety of ecosystem functions. However, to date the majority of soil microbial ecology studies target phylogenetic rRNA markers, while a smaller number target functional markers linked to soil processes. We validated a new primer set targeting citrate synthase (gtlA), a central enzyme in the citric acid cycle linked to aerobic respiration. Primers for a 225bp fragment suitable for qPCR were tested for specificity and assay performance verified on multiple soils. Clone libraries of the PCR-amplified gtlA gene exhibited high diversity and recovered most major groups identified in a previous 16S rRNA gene study. Comparisons among bacterial communities based on gtlA sequencing using UniFrac revealed differences among the experimental soils studied. Conditions for gtlA qPCR were optimized and calibration curves were highly linear (R 2 >0.99) over six orders of magnitude (4.56×105 to 4.56×1011 copies), with high amplification efficiencies (>1.7). We examined the performance of the gtlA qPCR across a variety of soils and ecosystems, spanning forests, old fields and agricultural areas. We were able to amplify gtlA genes in all tested soils, and detected differences in gtlA abundance within and among environments. These results indicate that a fully developed gtlA-targeted qPCR approach may have potential to link microbial community characteristics with changes in soil respiration. [Copyright &y& Elsevier]

Functional markers reduce the risk of false selection in marker-assisted selection (MAS), and recombination between the molecular marker and the gene of interest. The present study was conducted with the objective of combining the important basmati quality traits with resistance to bacterial leaf blight (BLB) by a combination of phenotypic selection and MAS. Screening of 29 recombinants for BLB resistant genes along with aroma ( fgr) gene was performed using a suite of different markers. Twenty genotypes were found in homozygous condition for Xa21 and xa13 genes. Restriction digestion analysis with BsrI, gave nine resistant recombinants. Out of nine resistant recombinants, seven were in homozygous and two were in heterozygous conditions. Recombinants viz., IRS 5441-2-21, IRS 5441-2-79, IRS 5441-2-81, IRS 5441-2-85 and IRS 5441-2-91 possessed all the three BLB resistance genes and fgr in the homozygous condition. Recombinants with enhanced resistance to BLB, basmati quality and desirable agronomic traits were identified. This germplasm can be directly developed into commercial varieties or used as immediate potential donors of BLB resistance in basmati breeding programs. [ABSTRACT FROM AUTHOR]

Development of high-yielding cereal crops could meet increasing global demands for food, feed and bio-fuels. Wheat is one of the world's most important cereal crops. The biosynthesis of starch is the major determinant of yield in wheat. Two starch biosynthesis genes, the waxy ( Wx) genes and the starch synthase IIa ( SSIIa) genes, were amplified and sequenced in 92 diverse wheat genotypes using genome-specific primers. Nucleotide diversity, haplotype analysis and association mapping were performed. The first exon (5′-UTR) and the first intron of the three homoeologous Wx genes were isolated using expressed sequence tag sequences. The Wx genes contained 12 exons separated by 11 introns. SNP (single nucleotide polymorphism) frequency ranged from 1 SNP/3,648 bp for Wx- D1 to 1 SNP/135 bp for SSIIa- A1, with an average of 1 SNP/230 bp. The average SNP frequencies in exon and intron regions were 1 SNP/322 bp and 1 SNP/228 bp, respectively. Thirty, 23 and 5 SNPs were identified and formed five, six and five haplotypes for SSIIa- A1, SSIIa- B1 and SSIIa- D1, respectively. However, no association was found between these SNPs and seven yield-related traits. Twenty-two, 15 and 1 SNPs were detected and formed nine, five and two haplotypes for Wx- A1, Wx- B1 and Wx- D1, respectively. Three unique nucleotides C+A+T at SNP5, SNP6 and SNP12 formed Wx-B1-H3, which was significantly associated with increased grain weight, thousand kernel weight, and total starch content in three spring wheat genotypes and five winter wheat genotypes. Cost-effective and co-dominant SNP markers were developed using temperature-switch (TS)-PCR and are being used for marker-assisted selection of doubled haploid lines with enhanced grain yield and starch content in winter wheat breeding programs. [ABSTRACT FROM AUTHOR]

The melon gene Fom- 2, which confers resistance to Fusarium oxysporum f.sp. melonis (Fom) races 0 and 1, has been previously characterized by map-based cloning, and it encodes a protein with a nucleotide binding site (NBS) and leucine-rich repeats (LRRs). Here, we used the primer Fom2-LRR to clone and sequence a partial LRR region of the Fom- 2 gene in 11 melon accessions resistant to Fusarium wilt from various geographic regions. Our work revealed that the structure of the partial LRR domain is highly conserved between eight of these resistant accessions and is similar to the resistant allele in the previously characterized PI-161375 line. Conversely, PI-124111 is a unique line that presents the same resistant allele that was previously described in the MR-1 line. The accession Cum-355 presents a protein that differs from that encoded by both the resistant lines PI-161375 and MR-1. This result suggests that Cum-355 has a new resistant allele of Fom- 2 that determines the same specificity. Importantly, based on the sequence of the Fom-2 LRR domain, two sequence characterized amplified region (SCAR) markers, Fom2-R and Fom2-S, were developed for Fom- 2 resistant and susceptible alleles, respectively. These allele-specific PCR markers could be used as co-dominant markers when their primer pairs were combined in a multiplex PCR reaction. The specificity of these functional markers (FM) was validated on a set of 27 genotypes representing several melon types. These FM markers are expected to enhance the reliability and cost-effectiveness of marker-assisted selection for the Fom- 2 gene in melon. [ABSTRACT FROM AUTHOR]

Grain yield is one of the most important indexes in rice breeding, which is governed by quantitative trait loci (QTLs). Different mapping populations have been used to explore the QTLs controlling yield related traits. Primary populations such as F2 and recombinant inbred line populations have been widely used to discover QTLs in rice genome-wide, with hundreds of yield-related QTLs detected. Advanced populations such as near isogenic lines (NILs) are efficient to further fine-map and clone target QTLs. NILs for primarily identified QTLs have been proposed and confirmed to be the ideal population for map-based cloning. To date, 20 QTLs directly affecting rice grain yield and its components have been cloned with NIL-F2 populations, and 14 new grain yield QTLs have been validated in the NILs. The molecular mechanisms of a continuously increasing number of genes are being unveiled, which aids in the understanding of the formation of grain yield. Favorable alleles for rice breeding have been 'mined' from natural cultivars and wild rice by association analysis of known functional genes with target trait performance. Reasonable combination of favorable alleles has the potential to increase grain yield via use of functional marker assisted selection. [ABSTRACT FROM AUTHOR]

Grain yield is one of the most important indexes in rice breeding, which is governed by quantitative trait loci (QTLs). Different mapping populations have been used to explore the QTLs controlling yield related traits. Primary populations such as F2 and recombinant inbred line populations have been widely used to discover QTLs in rice genome-wide, with hundreds of yield-related QTLs detected. Advanced populations such as near isogenic lines (NILs) are efficient to further fine-map and clone target QTLs. NILs for primarily identified QTLs have been proposed and confirmed to be the ideal population for map-based cloning. To date, 20 QTLs directly affecting rice grain yield and its components have been cloned with NIL-F2 populations, and 14 new grain yield QTLs have been validated in the NILs. The molecular mechanisms of a continuously increasing number of genes are being unveiled, which aids in the understanding of the formation of grain yield. Favorable alleles for rice breeding have been 'mined' from natural cultivars and wild rice by association analysis of known functional genes with target trait performance. Reasonable combination of favorable alleles has the potential to increase grain yield via use of functional marker assisted selection. [ABSTRACT FROM AUTHOR]

Jute is an important natural fibre crop, which is only second to cotton in its importance at the global level. It is mostly grown in Indian subcontinent and has been recently used for the development of genomics resources. We recently initiated a programme to develop simple sequence repeat markers and reported a set of 2469 SSR that were developed using four SSR-enriched libraries (Mir et al.). In this communication, we report an additional set of 607 novel SSR in 393 SSR containing sequences. However, primers could be designed for only 417 potentially useful SSR. Polymorphism survey was carried out for 374 primer pairs using two parental genotypes (JRO 524 and PPO4) of a mapping population developed for fibre fineness; only 66 SSR were polymorphic. Owing to a low level of polymorphism between the parental genotypes and a high degree of segregation distortion in recombinant inbred lines, genotypic data of only 53 polymorphic SSR on the mapping population consisting of 120 RIL could be used for the construction of a linkage map; 36 SSR loci were mapped on six linkage groups that covered a total genetic distance of 784.3 cM. Hopefully, this map will be enriched with more SSR loci in future and will prove useful for identification of quantitative trait loci/genes for molecular breeding involving improvement of fibre fineness and other related traits in jute. [ABSTRACT FROM AUTHOR]

Background: Functional genetic markers have important implications for genetic analysis by providing direct estimation of functional diversity. Although high throughput sequencing techniques for functional diversity analysis are being developed nowadays, the use of already well established variable markers present in candidate genes is still an interesting alternative for mapping purposes and functional diversity studies. SSR markers are routinely used in most plant and animal breeding programs for many species including Eucalyptus. SSR markers derived from candidate genes (SSR-CG) can be used effectively in co-segregation studies and marker-assisted diversity management. Results: In the present study, eight new non reported SSRs were identified in seven candidate genes for wood properties in Eucalyptus globulus: cinnamoyl CoA reductase (CCR), homocysteine S-methyltransferase (HMT), shikimate kinase (SK), xyloglucan endotransglycosylase 2 (XTH2), cellulose synthase 3 (CesA3), glutathione S-transferase (GST) and the transcription factor LIM1. Microsatellites were located in promoters, introns and exons, being most of them CT dinucleotide repeats. Genetic diversity of these eight CG-derived SSR-markers was explored in 54 unrelated genotypes. Except for XTH2, high levels of polymorphism were detected: 93 alleles (mean of 13.1 sd 1.6 alleles per locus), a mean effective number of alleles (Ne) of 5.4 (sd 1.6), polymorphic information content values (PIC) from 0.617 to 0.855 and probability of Identity (PI) ranging from 0.030 to 0.151. Conclusions: This is the first report on the identification, characterization and diversity analysis of microsatellite markers located inside wood quality candidate genes (CG) from Eucalyptus globulus. This set of markers is then appropriate for characterizing genetic variation, with potential usefulness for quantitative trait loci (QTL) mapping in different eucalypts genetic pedigrees and other applications such as fingerprinting and marker assisted diversity management. [ABSTRACT FROM AUTHOR]

With 1 figure and 3 tables The Rosodae supertribe is composed of economically important species (strawberry, rose and raspberry). In order to extend the repertoire of markers for optimizing genetic mapping and synteny within this supertribe, we developed 20 functional markers from two Fragaria cDNA libraries enriched in AG/AC microsatellite motifs. We tested their transferability within the Rosodae supertribe, including two reference diploid and octoploid strawberry segregating populations. Out of these 20 functional markers, 70% revealed polymorphism in both diploid and octoploid Fragaria segregating populations and were mapped on the diploid genetic map to provide useful information for further marker‐assisted selection. These 20 EST‐SSRs were also tested in 17 wild Fragaria species and displayed a high cross‐species transferability (mean of 95%), providing new markers for further diversity studies within the genus. These markers were also transferable to the phylogenetically close Potentilla genus, whereas their transferability was lower for the polyploid Duchesnea indica species and for the more distant Rosa genus. Their use for genetic mapping within the Rosodae will further permit a comparison of genetic information among the different genera. [ABSTRACT FROM AUTHOR]

Three single nucleotide polymorphism (SNP) sites in which amino acids had changed were detected by sequence analysis within the leucine-rich repeat (LRR) region of the Fom- 2 gene. Cleaved amplified polymorphic sequence (CAPS) and allele-specific PCR (AS-PCR) methods were employed to explore the SNP validation linked to fusarium wilt resistance in the F and F generations simultaneously. Homozygous- and heterozygous-resistant genotypes and homozygous-susceptible genotype could be clearly distinguished using the CAPS method, and three detected SNP sites were observed to be linked to fusarium wilt resistance, with a segregation ratio of 1:2:1 in the F generation. In addition, heterozygous-resistant and homozygous-susceptible genotypes could be clearly distinguished in the F generation using the AS-PCR method, showing a 3:1 segregation in terms of resistant and susceptible genotypes in the F generation. We therefore developed SNP-based functional markers (FMs) and identified some melon germplasm resistant to fusarium wilt by FM analysis within melon species. In conclusion, the SNP-based FMs originating from the SNP site of the Fom- 2 LRR region were determined to be linked to fusarium wilt resistance and showed promise in the enhancement of breeding in melon. [ABSTRACT FROM AUTHOR]

Cardiac progenitor cells (CPCs), migrating from heart tissue, in culture aggregate to form cardiospheres (CSs) in which replication and cardiogenic differentiation occur. However, the frequency of functional differentiation in CSs and the role of cell clustering in supporting it remain to be established. The aim of our study is to quantify differentiation of a muscle-type Ca release mechanism in CS-derived cells, correlate it with cardiac differentiation markers and test its dependency on CS formation. CPCs migrating from murine cardiac explants were studied prior and after CSs formation (Pre-CS and Post-CS). Inducibility of RyR- and IP3-R-mediated Ca transients in individual cells was tested by exposure to caffeine and ATP, respectively; expression of cardiac and non-cardiac lineage markers was assessed. Caffeine responsiveness was negligible in Pre-CS cells and increased by 7.5 fold in Post-CS cells (3.6 vs. 26.9%; p < 0.05), and was closely correlated with activation of the cardiac TnI gene promoter. ATP-induced responses, frequent in Pre-CS (86%), were slightly increased in Post-CS cells (94%; p < 0.05). Expression of cardiac-specific Ca-handling proteins (Cav1.2, NCX1, RyR2, SERCA2a) was either limited to the Post-CS stage, or markedly enhanced. CS beating was infrequent, but its pharmacology was compatible with cardiac excitation-contraction coupling. Expression of non-cardiac lineage was low in general, and similar between Pre- and Post-CS cells. Culture conditions inhibiting CSs formation prevented the increase in caffeine responders. In conclusion, clustering in CSs leads to the induction of a muscle-specific functional response in about 30% of CPCs; this is accompanied by development of a cardiac-specific expression pattern. [ABSTRACT FROM AUTHOR]

With 1 figure and 3 tables Evolving high yielding rice genotypes with durable resistance to bacterial leaf blight (BLB) is pertinent considering the extensive damage caused by the disease in most of the rice growing regions. Two high yielding BLB susceptible indica rice cultivars, ‘ADT43’ and ‘ASD16’ popular among farmers and consumers across South India have been introgressed with three BLB resistance genes xa5, xa13 and Xa21 from isoline IRBB60 using functional markers. The F2 populations of 500 plants from ADT43 × IRBB60 and 806 plants from ASD16 × IRBB60 crosses were screened for the presence of all the three resistance genes. Thirty genotypes with three genes in homozygous and 55 genotypes with two genes in homozygous and one gene in heterozygous condition were identified. These pyramided genotypes with two or three resistance genes exhibited high levels of resistance against two predominant Xanthomonas oryzae isolates of South India. Among the 30 pyramided genotypes ( xa5 + xa13 + Xa21), 12 were found to be significantly high yielding with desirable agronomic characteristics and the selection efficiency of the present markers was hundred percent. [ABSTRACT FROM AUTHOR]

Target region amplification polymorphism (TRAP) markers were used to estimate the genetic similarity (GS) among 53 sugarcane varieties and five species of the Saccharum complex. Seven fixed primers designed from candidate genes involved in sucrose metabolism and three from those involved in drought response metabolism were used in combination with three arbitrary primers. The clustering of the genotypes for sucrose metabolism and drought response were similar, but the GS based on Jaccard's coefficient changed. The GS based on polymorphism in sucrose genes estimated in a set of 46 Brazilian varieties, all of which belong to the three Brazilian breeding programs, ranged from 0.52 to 0.9, and that based on drought data ranged from 0.44 to 0.95. The results suggest that genetic variability in the evaluated genes was lower in the sucrose metabolism genes than in the drought response metabolism ones. [ABSTRACT FROM AUTHOR]

Expressed sequence tags (ESTs) offer the opportunity to exploit single, low-copy, conserved sequence motifs for the development of simple sequence repeats (SSRs). The authors have examined the Sugarcane Expressed Sequence Tag database for the presence of SSRs. To test the utility of EST-derived SSR markers, a total of 342 EST-SSRs, which represent a subset of over 2005 SSR-containing sequences that were located in the sugarcane EST database, could be designed from the nonredundant SSR-positive ESTs for possible use as potential genic markers. These EST-SSR markers were used to screen 18 sugarcane (Saccharum spp.) varieties. A high proportion (65.5%) of the above EST-SSRs, which gave amplified fragments of foreseen size, detected polymorphism. The number of alleles ranged from 2 to 24 with an average of 7.55 alleles per locus, while polymorphism information content values ranged from 0.16 to 0.94, with an average of 0.73. The ability of each set of EST-SSR markers to discriminate between varieties was generally higher than the polymorphism information content analysis. When tested for functionality, 82.1% of these 224 EST-SSRs were found to be functional, showing homology to known genes. As the EST-SSRs are within the expressed portion of the genome, they are likely to be associated to a particular gene of interest, improving their utility for genetic mapping; identification of quantitative trait loci, and comparative genomics studies of sugarcane. The development of new EST-SSR markers will have important implications for the genetic analysis and exploitation of the genetic resources of sugarcane and related species and will provide a more direct estimate of functional diversity. Les étiquettes de séquences exprimées (EST) offrent une opportunité d’exploiter des motifs conservés présents en simple ou peu de copies pour le développement de microsatellites (SSR). Les auteurs ont inspecté la base de données d’EST de la canne à sucre pour la présence de SSR. Pour vérifier l’utilité des SSR dérivés d’EST, 342 EST-SSR, lesquels représentent un sous-ensemble des 2 005 séquences contenant des SSR qui ont été trouvées au sein de cette banque d’EST de la canne à sucre, étaient propices au développement de marqueurs génétiques parmi les EST non-redondants contenant des EST. Ces marqueurs EST-SSR ont été employés pour cribler 18 variétés de canne à sucre (Saccharum spp.). Une proportion élevée (65,5%) de ces EST-SSR qui ont amplifié des produits de la taille attendue étaient polymorphes. Le nombre d’allèles variait entre 2 et 24, pour une moyenne de 7,55 allèles par locus, et les valeurs de contenu d’information de polymorphisme variaient entre 0,16 et 0,94, pour une moyenne de 0,73. La capacité de tels jeux de marqueurs à discriminer entre les variétés était généralement plus élevée que l’analyse de contenu d’information de polymorphisme. Lorsque testés pour leur fonction, 82,1% de ces 224 marqueurs EST-SSR se sont avérés fonctionnels en montrant de l’homologie avec des gènes connus. Puisque ces EST-SSR font partie de la portion exprimée du génome, ils sont vraisemblablement associés à des gènes d’intérêt, ce qui augmente leur utilité pour des fins de cartographie génétique, pour l’identification de locus de caractères quantitatifs et pour des études de génomique comparative chez la canne à sucre. Le développement de nouveaux marqueurs EST-SSR aura d’importantes retombées en matière d’analyse génétique et d’exploitation des ressources génétiques chez la canne à sucre et des espèces apparentées en plus de fournir un estimé plus direct de la diversité fonctionnelle. [ABSTRACT FROM AUTHOR]

Speed and cost–effective techniques for evaluation of plant materials to provide information before entering the next cycle of selection are critical for success in plant breeding. Whether or not a ‘new’ technique realizes its potential depends on technical and economic considerations. Biotechnology-based research tools such as doubled haploid technology and molecular markers have already demonstrated their value for application in plant breeding. In the area of genomics, implementation of functional markers (FMs) is currently of particular interest. The pipeline from plant materials to FM data points for any application includes maceration of plant material, DNA isolation and sample preparation. For each step of this pipeline, a number of techniques are available, and no single method is ideally suited for all applications. The challenge is to meet the needs of many different scenarios which are present in a modern breeding programme such as the use of (1) few markers for genotyping hundreds of samples (e.g., marker assisted backcrossing (MAB)), (2) few markers in thousands of samples (e.g., screening for GMO), (3) hundreds to thousands of markers for hundreds to thousands of samples (e.g., genetic characterization of breeding materials (fingerprinting)). This paper compares different techniques for each of the steps from plant material to FM data point, with the main emphasis on SNP detection platforms, assuming that multiple FMs will become available in the near future. We focus on technical and economic aspects and discuss which techniques are most suitable for each of the scenarios using wheat as a model. [ABSTRACT FROM AUTHOR]

Low-molecular-weight glutenin subunits (LMW-GS) have great effect on wheat processing quality, but were numerous and difficult to dissect by SDS-PAGE. The development of functional markers may be the most effective way for a clear discrimination of different LMW-GS genes. In the present study, three different approaches were used to identify SNPs of different genes at Glu-D3 and Glu-B3 loci in bread wheat for the development of six STS markers (3 for Glu-D3 and 3 for Glu-B3 genes) that were validated with distinguished wheat cultivars. Firstly, seven LMW-GS gene sequences ( AY585350, AY585354, AY585355, AY585356, AY585349, AY585351 and AY585353 ) from Aegilops tauschii, the diploid donor of the D-genome of bread wheat, were chosen to design seven pairs of AS-PCR primers for Glu-D3 genes. By amplifying the corresponding genes from five bread wheat cultivars with different Glu-D3 alleles ( a, b, c, d and e) and Ae. tauschii, a primer set, S13F2/S13R1, specific to the gene AY585356, was found to be positive to cultivars with alleles Glu-D3c and d. Nevertheless, the other five pairs of primers designed from AY585350, AY585349, AY585353, AY585354 and AY585355, respectively, did not produce specific PCR products to the cultivars tested. Secondly, all the PCR products from the five primer sets without specific characteristics were sequenced and an SNP from the gene AY585350 was detected in the cultivar Hartog, which resulted in the second STS marker S1F1/S1R3 specific to the allelic variant of AY585350. Thirdly, three Glu-D3 sequences ( AB062851, AB062865 and AB062872) and three Glu-B3 sequences ( AB062852, AB062853 and AB062860) defined by Ikeda et al. () were chosen to query wheat EST and NR databases, and DNA markers were developed based on the putative SNPs among the sequences. Using this approach, four STS markers were developed and validated with 16-19 bread wheat cultivars. The primer set T1F4/T1R1 was also a Glu-D3 gene-specific marker for AB062872, while T2F2/T2R2, T5F3/T5R1 and T13F4/T13R3 were all Glu-B3 gene specific markers for AB062852, BF293671 and AY831800, respectively. The chromosomal locations of the six markers were verified by amplifying the genomic DNA of Ae. tauschii (DD), T. monococcum (AA) and T. turgidum (AABB) entries, as well as Chinese Spring and its group 1 chromosome nulli-tetrasomic lines. The results are useful to discriminate the corresponding Glu-D3 and Glu-B3 genes in wheat breeding programs. [ABSTRACT FROM AUTHOR]

Human-induced changes in land use lead to major changes in plant community composition which have strong effects on ecosystem processes. Here, we tested the hypothesis that changes in traits of living plants induced by such changes resulted in changes in the quality and decay properties of the litter produced by the different communities. This was done in the context of a secondary succession following land abandonment in the Mediterranean region of Southern France. During the course of succession, species with high specific leaf area (the ratio of leaf area to leaf mass), low leaf dry matter content (the ratio of leaf dry mass to leaf fresh mass) and high leaf nitrogen concentration were progressively replaced by species with opposite characteristics. Accordingly, the initial litter concentrations of carbon (C) and nitrogen (N) decreased, while their C:N ratio and their hemicellulose concentration increased with time after abandonment. Early-successional communities had faster rates of litter decay and N release from litter, but these differences damped out with decomposition time. Nitrogen release from litter was related to initial litter chemical composition, particularly to its N concentration. This also held for litter decay rate, but only during the first 18 months of decomposition. Community functional parameters (i.e. trait values weighed according to the relative abundance of species) were tightly linked to initial litter N concentration, and thereby to litter decay and N loss rates. The strongest correlations were found with leaf dry matter content, which therefore appears as a powerful marker of litter properties. This provides further evidence that characteristics of living leaves persist in litter, and that some ecosystem processes can be inferred from plant functional traits. [ABSTRACT FROM AUTHOR]

Contents Summary 719 I. Introduction 720 II. General methodology 721 III. Methodological considerations 722 IV. Application of microarrays in the soil environment 726 V. Conclusions 731 Acknowledgements 732 References 732 Soil microbial communities are responsible for important physiological and metabolic processes. In the last decade soil microorganisms have been frequently analysed by cultivation-independent techniques because only a minority of the natural microbial communities are accessible by cultivation. Cultivation-independent community analyses have revolutionized our understanding of soil microbial diversity and population dynamics. Nevertheless, many methods are still laborious and time-consuming, and high-throughput methods have to be applied in order to understand population shifts at a finer level and to be better able to link microbial diversity with ecosystems functioning. Microbial diagnostic microarrays (MDMs) represent a powerful tool for the parallel, high-throughput identification of many microorganisms. Three categories of MDMs have been defined based on the nature of the probe and target molecules used: phylogenetic oligonucleotide microarrays with short oligonucleotides against a phylogenetic marker gene; functional gene arrays containing probes targeting genes encoding specific functions; and community genome arrays employing whole genomes as probes. In this review, important methodological developments relevant to the application of the different types of diagnostic microarrays in soil ecology will be addressed and new approaches, needs and future directions will be identified, which might lead to a better insight into the functional activities of soil microbial communities. [ABSTRACT FROM AUTHOR]

Functional markers (FMs) are derived from polymorphic sites within genes causally involved in phenotypic trait variation (Andersen, J.R. & T. Lübberstedt, 2003. Trends Plant Sci 8: 554–560). FM development requires allele sequences of functionally characterized genes from which polymorphic, functional motifs affecting plant phenotype can be identified. In maize and other species with low levels of linkage disequilibrium, association studies have the potential to identify sequence motifs, such as a few nucleotides or insertions/deletions, affecting trait expression. In one of the pioneering studies, nine sequence motifs in the dwarf8 gene of maize were shown to be associated with variation for flowering time (Thornsberry, J.M., M.M. Goodman, J. Doebley, S. Kresovich, D. Nielsen & E.S. Buckler, 2001. Nat Genet 28: 286–289). Proof of sequence motif function can be obtained by comparing isogenic genotypes differing in single sequence motifs. At current, the most appropriate approach for this purpose in crops is targeting induced local lesions in genomes (TILLING) (McCallum, C.M., L. Comai, E.A. Greene & S. Henikoff, 2000. Nat Biotechnol 18: 455–457). In central Europe, maize is mainly grown as forage crop, with forage quality as major trait, which can be determined as proportion of digestible neutral detergent fiber (DNDF). Brown midrib gene knock out mutations have been shown to be beneficial for forage quality but disadvantageous for overall agronomic performance. Two brown midrib genes ( bm1 and bm3) have been shown to be involved in monolignol biosynthesis. These two and additional lignin biosynthesis genes have been isolated based on sequence homology. Additional candidate genes putatively affecting forage quality have been identified by expression profiling using, e.g., isogenic bm lines. Furthermore, we identified an association between a polymorphism at the COMT locus and DNDF in a collection of European elite inbred lines. [ABSTRACT FROM AUTHOR]

Abstract Public genomic databases have provided new directions for molecular marker development and initiated a shift in the types of PCR-based techniques commonly used in plant science. Alongside commonly used arbitrarily amplified DNA markers, other methods have been developed. Targeted fingerprinting marker techniques are based on the well-established practices of arbitrarily amplified DNA methods, but employ novel methodological innovations such as the incorporation of gene or promoter elements in the primers. These markers provide good reproducibility and increased resolution by the concurrent incidence of dominant and co-dominant bands. Despite their promising features, these semi-random markers suffer from possible problems of collision and non-homology analogous to those found with randomly generated fingerprints. Transposable elements, present in abundance in plant genomes, may also be used to generate fingerprints. These markers provide increased genomic coverage by utilizing specific targeted sites and produce bands that mostly seem to be homologous. The biggest drawback with most of these techniques is that prior genomic information about retrotransposons is needed for primer design, prohibiting universal applications. Another class of recently developed methods exploits length polymorphism present in arrays of multi-copy gene families such as cytochrome P450 and β-tubulin genes to provide cross-species amplification and transferability. A specific class of marker makes use of common features of plant resistance genes to generate bands linked to a given phenotype, or to reveal genetic diversity. Conserved DNA-based strategies have limited genome coverage and may fail to reveal genetic diversity, while resistance genes may be under specific evolutionary selection. Markers may also be generated from functional and/or transcribed regions of the genome using different gene-targeting approaches coupled with the use of RNA information. Such techniques have the potential to generate phenotypically linked functional markers, especially when fingerprints are generated from the transcribed or expressed region of the genome. It is to be expected that these recently developed techniques will generate larger datasets, but their shortcomings should also be acknowledged and carefully investigated.

In the past, a major challenge for nutrition research was in defining indicators of nutritional adequacy. More recently, the research base related to the role of nutrition in chronic disease has expanded sufficiently to permit moving beyond deficiency indicators to other indicators with broader functional significance. Thus, nutrition research is faced with the new challenge of defining ‘optimal nutrition’. One definition of optimal nutrition with respect to any particular nutrient could be when a functional marker reaches an ‘optimal value’ or plateau beyond which it is no longer affected by intake or stores of the nutrient. A functional marker of nutrient status could be defined as a physiological or biochemical factor which (1) is related to function or effect of the nutrient in target tissue(s) and (2) is affected by dietary intake or stores of the nutrient (which may include markers of disease risk). Examples of such indicators or markers are those related to risk of chronic diseases such as osteoporosis, CHD, or hypertension. The present review focuses on the concept of optimal nutrition with respect to three nutrients, Ca, Mg and P. However, for P and Mg there are as yet no functional indicators which respond to dietary intake, and in such cases nutrient requirements are established using more traditional approaches, e.g. balance data. For Ca, there has been interest in using maximal Ca retention, which is based on balance data, bone mass measurements and biomarkers of bone turnover as useful functional indicators of the adequacy of Ca intake. [ABSTRACT FROM PUBLISHER]

Background: Cognitive impairment is associated with long-term disability that results in the deterioration of both the social and professional status of individuals with schizophrenia. The impact of antipsychotic therapy on cognitive function is insufficient. Cognitive training is therefore proposed as a tool for cognitive rehabilitation in schizophrenia. In this study we investigated the effect of self-administered cognitive training using a smartphone-based application on the cognitive function of paranoid schizophrenia patients focusing on response time, correct answer rate, incorrect answer rate, and fatigability to check, if these functions can be functional markers of successful cognitive-smartphone rehabilitation. Methods: 1-year multicenter, open-label randomized study was conducted on 290 patients in a state of symptomatic remission. 191 patients were equipped with the full version of the application and conducted cognitive training twice a week. Reference group (n = 99) was provided with a version of the application having only limited functionality, testing the cognitive performance of patients every 6 months. Results: Statistically significant improvement was observed in both the rate of correct answers (by 4.8%, p = 0.0001), and cognitive fatigability (by 2.9%, p = 0.0001) in the study group, along with a slight improvement in the rate of incorrect answers (by 0.9%, p = 0.15). In contrast, the reference group, who performed cognitive training every 6 months, demonstrated no significant changes in any cognitive activities. Conclusions: Cognitive trainings facilitated by a smartphone-based application, performed regularly for a longer period of time are feasible and may have the potential to improve the cognitive functioning of individuals with schizophrenia. Correct answers and cognitive fatigability have potential to be functional markers of successful smartphone-based psychiatric rehabilitations in schizophrenia patients. [ABSTRACT FROM AUTHOR]

Background: The species Olea europaea includes cultivated varieties (subsp. europaea var. europaea), wild plants (subsp. europaea var. sylvestris), and five other subspecies spread over almost all continents. Single nucleotide polymorphisms in the expressed sequence tag able to underline intra-species differentiation are not yet identified, beyond a few plastidial markers. Methods: In the present work, more than 1000 transcript-specific SNP markers obtained by the genotyping of 260 individuals were studied. These genotypes included cultivated, oleasters, and samples of subspecies guanchica, and were analyzed in silico, in order to identify polymorphisms on key genes distinguishing different Olea europaea forms. Results: Phylogeny inference and principal coordinate analysis allowed to detect two distinct clusters, clearly separating wilds and guanchica samples from cultivated olives, meanwhile the structure analysis made possible to differentiate these three groups. Sequences carrying the polymorphisms that distinguished wild and cultivated olives were analyzed and annotated, allowing to identify 124 candidate genes that have a functional role in flower development, stress response, or involvement in important metabolic pathways. Signatures of selection that occurred during olive domestication, were detected and reported. Conclusion: This deep EST–SNP analysis provided important information on the genetic and genomic diversity of the olive complex, opening new opportunities to detect gene polymorphisms with potential functional and evolutionary roles, and to apply them in genomics-assisted breeding, highlighting the importance of olive germplasm conservation. [ABSTRACT FROM AUTHOR]

Advances in molecular biology including genomics, high-throughput sequencing, and genome editing enable increasingly faster and more precise cultivar development. Identifying genes and functional markers (FMs) that are highly associated with plant phenotypic variation is a grand challenge. Functional genomics approaches such as transcriptomics, targeting induced local lesions in genomes (TILLING), homologous recombinant (HR), association mapping, and allele mining are all strategies to identify FMs for breeding goals, such as agronomic traits and biotic and abiotic stress resistance. The advantage of FMs over other markers used in plant breeding is the close genomic association of an FM with a phenotype. Thereby, FMs may facilitate the direct selection of genes associated with phenotypic traits, which serves to increase selection efficiencies to develop varieties. Herein, we review the latest methods in FM development and how FMs are being used in precision breeding for agronomic and quality traits as well as in breeding for biotic and abiotic stress resistance using marker assisted selection (MAS) methods. In summary, this article describes the use of FMs in breeding for development of elite crop cultivars to enhance global food security goals. [ABSTRACT FROM AUTHOR]

The use of natural marine bacteria as "oil sensors" for the detection of pollution events can be suggested as a novel way of monitoring oil occurrence at sea. Nucleic acid-based devices generically called genosensors are emerging as potentially promising tools for in situ detection of specific microbial marker genes suited for that purpose. Functional marker genes are particularly interesting as targets for oil-related genosensing but their identification remains a challenge. Here, seawater samples, collected in tanks with oil addition mimicking a realistic oil spill scenario, were filtered and archived by the Environmental Sample Processor (ESP), a fully robotized genosensor, and the samples were then used for post-retrieval metatranscriptomic analysis. After extraction, RNA from ESP-archived samples at start, Day 4 and Day 7 of the experiment was used for sequencing. Metatranscriptomics revealed that several KEGG pathways were significantly enriched in samples exposed to oil. However, these pathways were highly expressed also in the non-oil-exposed water samples, most likely as a result of the release of natural organic matter from decaying phytoplankton. Temporary peaks of aliphatic alcohol and aldehyde dehydrogenases and monoaromatic ring-degrading enzymes (e.g., ben, box, and dmp clusters) were observed on Day 4 in both control and oil-exposed and non-exposed tanks. Few alkane 1-monooxygenase genes were upregulated on oil, mostly transcribed by families Porticoccaceae and Rhodobacteraceae, together with aromatic ring-hydroxylating dioxygenases, mostly transcribed by Rhodobacteraceae. Few transcripts from obligate hydrocarbonoclastic genera of Alcanivorax, Oleispira and Cycloclasticus were significantly enriched in the oil-treated exposed tank in comparison to control the non-exposed tank, and these were mostly transporters and genes involved in nitrogen and phosphorous acquisition. This study highlights the importance of seasonality, i.e., phytoplankton occurrence and senescence leading to organic compound release which can be used preferentially by bacteria over oil compounds, delaying the latter process. As a result, such seasonal effect can reduce the sensitivity of genosensing tools employing bacterial functional genes to sense oil. A better understanding of the use of natural organic matter by bacteria involved in oil-biodegradation is needed to develop an array of functional markers enabling the rapid and specific in situ detection of anthropogenic pollution. [ABSTRACT FROM AUTHOR]

Regulatory T cells (Tregs) are enriched in the tumor microenvironment and play key roles in immune evasion of cancer cells. Cell surface markers specific for tumor-infiltrating Tregs (TI-Tregs) can be effectively targeted to enhance antitumor immunity and used for stratification of immunotherapy outcomes. Here, we present a systems biology approach to identify functional cell surface markers for TI-Tregs. We selected differentially expressed genes for surface proteins of TI-Tregs and compared these with other CD4 + T cells using bulk RNA-sequencing data from murine lung cancer models. Thereafter, we filtered for human orthologues with conserved expression in TI-Tregs using single-cell transcriptome data from patients with non-small cell lung cancer (NSCLC). To evaluate the functional importance of expression-based markers of TI-Tregs, we utilized network-based measure of context-associated centrality in a Treg-specific coregulatory network. We identified TNFRSF9 (also known as 4-1BB or CD137 ), a previously reported target for enhancing antitumor immunity, among the final candidates for TI-Treg markers with high functional importance score. We found that the low TNFRSF9 expression level in Tregs was associated with enhanced overall survival rate and response to anti-PD-1 immunotherapy in patients with NSCLC, proposing that TNFRSF9 promotes immune suppressive activity of Tregs in tumor. Collectively, these results demonstrated that integrative transcriptome and network analysis can facilitate the discovery of functional markers of tumor-specific immune cells to develop novel therapeutic targets and biomarkers for boosting cancer immunotherapy., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Author(s).)

Lentil is an important food legume crop that has large and complex genome. During past years, considerable attention has been given on the use of next generation sequencing for enriching the genomic resources including identification of SSR and SNP markers, development of unigenes, transcripts, and identification of candidate genes for biotic and abiotic stresses, analysis of genetic diversity and identification of genes/ QTLs for agronomically important traits. However, in other crops including pulses, next generation sequencing has revolutionized the genomic research and helped in genomic assisted breeding rapidly and cost effectively. The present review discuss current status and future prospects of the use NGS based breeding in lentil.

A total of 17,439 mature miRNAs (~ 21 nt) earlier generated through RNA seq in the pomegranate were used for in silico analysis. After complexity reduction, a total of 1922 representative mature miRNAs were selected and used as query sequences against pomegranate genome to retrieve 2540 homologous contigs with flanking regions (~ 800). By using pre-miRNA prediction web server, a total of 1028 true contigs harbouring pri-miRNAs encoding 1162 pre-miRNAs were identified. Survey of these sequences for SSRs yielded a total of 1358 and 238 SSRs specific to pri-miRNA and pre-miRNAs, respectively. Of these, primer pairs were designed for 897 pri-miRNA and 168 pre-miRNA SSRs. In pri-miRNA sequences, hexa-nucleotides repeats were found to be most abundant (44.18%) followed by mono- (18.41%) and di-nucleotide (17.01%), which is also observed in pre-miRNA sequences. Further, a set of 51 randomly selected pre-miRNA-SSRs was examined for marker polymorphism. The experimental validation of these markers on eight pomegranate genotypes demonstrated 92.15% polymorphism. Utility of these functional markers was confirmed via examination of genetic diversity of 18 pomegranate genotypes using 15 miRNA-SSRs. Further, potential application of miRNA-SSRs for discovery of trait specific candidate genes was showed by validating 51 mature miRNA against publically available 2047 EST sequences of pomegranate by target and network analysis. In summary, the current study offers novel functional molecular markers for pomegranate genetic improvement., (© Prof. H.S. Srivastava Foundation for Science and Society 2020.)