Plant Genomics

Background: Many molecular biology experiments start with DNA extraction. The quality and the quantity of the extracted DNA are very important for downstream analyses. Therefore, the DNA extraction process is very important and efforts should be aimed at constant improvement of the available protocols. Objective: The first objective of this study is to compare various lysing agents that use Sodium Dodecyl Sulphate (SDS) in DNA extraction process. The second objective is to report detailed protocol for the extraction of DNA from frozen animal sample. Methods: The efficiencies of four different lysing reagents involving SDS and a commercial lysing agent were compared. The reagents tested were ethylenediaminetetraacetic acid (EDTA), Sodium Chloride-Tris-EDTA (STE), Sodium chloride (NaCl) and tris acid (Tris). The four lysing agents investigated were SDS+EDTA+NaCl (SEN), SDS +EDTA+Tris (SET), SDS+STE+NaCl (SSN) and SDS+STE+Tris (STT). In addition, Qiagen (Q) lysing buffer was also inclu...

Halophytes are plants which naturally survive in saline environment. They account for ∼1% of the total flora of the world. They include both dicots and monocots and are distributed mainly in arid, semi-arid inlands and saline wet lands along the tropical and sub-tropical coasts. Salinity tolerance in halophytes depends on a set of ecological and physiological characteristics that allow them to grow and flourish in high saline conditions. The ability of halophytes to tolerate high salt is determined by the effective coordination between various physiological processes, metabolic pathways and protein or gene networks responsible for delivering salinity tolerance. The salinity responsive proteins belong to diverse functional classes such as photosynthesis, redox homeostasis; stress/defense, carbohydrate and energy metabolism, protein metabolism, signal transduction and membrane transport. The important metabolites which are involved in salt tolerance of halophytes are proline and proline analog (4-hydroxy-N-methyl proline), glycine betaine, pinitol, myo-inositol, mannitol, sorbitol, O-methylmucoinositol, and polyamines. In halophytes, the synthesis of specific proteins and osmotically active metabolites control ion and water flux and support scavenging of oxygen radicals under salt stress condition. The present review summarizes the salt tolerance mechanisms of halophytes by elucidating the recent studies that have focused on proteomic, metabolomic, and ionomic aspects of various halophytes in response to salinity. By integrating the information from halophytes and its comparison with glycophytes could give an overview of salt tolerance mechanisms in halophytes, thus laying down the pavement for development of salt tolerant crop plants through genetic modification and effective breeding strategies.

We are developing three alternative genotyping methods for marker-assisted selection (MAS) application in a collaborative project between IRRI, CIMMYT, and NARES partner institutions. Two methods, microarray-and FRET-based genotyping, are being developed as high-throughput and cost-effective techniques that could be utilized in regional laboratory hubs/core genotyping institutes while two other methods, dot blot-based genotyping (see accompanying poster) and PCR-ELISA, are being developed as low-cost approaches that can be adopted in laboratories with limited resources. We are validating these methods using previously developed markers and characterized or cloned bacterial blight resistance genes (xa5, Xa7, and Xa21) in rice. Funding from the Generation Challenge Program is gratefully acknowledged.

The Ilama is an important agricultural livestock in much of South America. The llama is increasing in popularity in the United States as a companion animal. Little work has been done to improve llama production using modern technology. A paucity of information is available regarding the llama genome. We report the construction of a llama bacterial artificial chromosome (BAC) library of about 196,224 clones in the vector pECBAC1. Using flow cytometry and bovine, human, mouse, and chicken as controls, we determined the llama genome size to be2.4×109 bp. The average insert size of the library is 137.8 kb corresponding to approximately 9-fold genome coverage. Further studies are needed to further characterize the library and llama genome. We anticipate that this new library will help facilitate future genomic studies in the llama.

Few public sector rice breeders have the capacity to use NGS-derived markers in their breeding programmes despite rapidly expanding repositories of rice genome sequence data. They rely on > 18,000 mapped microsatellites (SSRs) for marker-assisted selection (MAS) using gel analysis. Lack of knowledge about target SNP and InDel variant loci has hampered the uptake by many breeders of Kompetitive allele-specific PCR (KASP), a proprietary technology of LGC genomics that can distinguish alleles at variant loci. KASP is a cost-effective single-step genotyping technology, cheaper than SSRs and more flexible than genotyping by sequencing (GBS) or array-based genotyping when used in selection programmes. Before this study, there were 2015 rice KASP marker loci in the public domain, mainly identified by array-based screening, leaving large proportions of the rice genome with no KASP coverage. Here we have addressed the urgent need for a wide choice of appropriate rice KASP assays and demon...

Tomato is an important, popular, and versatile vegetable in the world and ranks number one in its contribution to the diet. One of the most common land degradation processes that affect agricultural production is soil salinization, however, organic production can be utilized to reduce the effect of salinity on many plants. The study aims to investigate the effect of different concentrations (0, 25, 31, 38, and 44%) of poultry manure (PM) on cherry tomato plants grown in marsh soil and to study the effectiveness of fertilizer in improving soil properties. The results showed that the application of PM in marsh soil increased plant height, root length, fresh and dry weight, the number of flowers and fruits and shoot potassium concentration, while shoot sodium concentration was decreased. The present study concluded that treating salt marsh soil with PM levels especially with PM4 level could reduce salinity stress damage on cherry tomato plants, increase biomass production and improve soil properties.

The concentration of copper (Cu), Zinc (Zn), manganese (Mn), lead (Pb), Chromium (Cr) and Nickel (Ni) in various vegetables Viz, Lettuce, Sesame and Okra irrigated from the bank of River Challawa Kano, Nigeria were assessed using atomic absorption spectrophotometer. The concentration ranges of Cu, Zn, Mn, Pb, Cr and Ni in the leaves of lettuce and sesame were found to be from 3.20-6.40, 10.00-35.00, 9.00-19.00, 6.00-13.00, 8.00-19.00 and 5.00-10.00mgkg -1 and

Abiotic stress induces several changes in plants at physiological and molecular level. Plants have evolved regulatory mechanisms guided towards establishment of stress tolerance in which epigenetic modifications play a pivotal role. We provide examples of gene expression changes that are brought about by conversion of active chromatin to silent heterochromatin and vice versa. Methylation of CG sites and specific modification of histone tail determine whether a particular locus is transcriptionally active or silent. We present a lucid review of epigenetic machinery and epigenetic alterations involving DNA methylation, histone tail modifications, chromatin remodeling, and RNA directed epigenetic changes.

Plants being sessile in nature encounter numerous biotic agents, including bacteria, fungi, viruses, insects, nematodes and protists. A great number of publications indicate that biotic agents significantly reduce crop productivity, although there are some biotic agents that symbiotically or synergistically co-exist with plants. Nonetheless, scientists have made significant advances in understanding the plant defence mechanisms expressed against biotic stresses. These mechanisms range from anatomy, physiology, biochemistry, genetics, development and evolution to their associated molecular dynamics. Using model plants, e.g., Arabidopsis and rice, efforts to understand these mechanisms have led to the identification of representative candidate genes, quantitative trait loci (QTLs), proteins and metabolites associated with plant defences against biotic stresses. However, there are drawbacks and insufficiencies in precisely deciphering and deploying these mechanisms, including only modest adaptability of some identified genes or QTLs to changing stress factors. Thus, more systematic efforts are needed to explore and expand the development of biotic stress resistant germplasm. In this chapter, we provided a comprehensive overview and discussed plant defence mechanisms involving molecular and cellular adaptation to biotic stresses. The latest achievements and perspective on plant molecular responses to biotic stresses, including gene expression, and targeted functional analyses of the genes expressed against biotic stresses have been presented and discussed.

Plants are frequently exposed to wide range of harsh environmental factors, such as drought, salinity, cold, heat, and insect attack. Being sessile in nature, plants have developed different strategies to adapt and grow under rapidly changing environments. These strategies involve rearrangements at the molecular level starting from transcription, regulation of mRNA processing, translation, and protein modification or its turnover. Plants show stress-specific regulation of transcription that affects their transcriptome under stress conditions. The transcriptionally regulated genes have different roles under stress response. Generally, seedling and reproductive stages are more susceptible to stress. Thus, stress response studies during these growth stages reveal novel differentially regulated genes or proteins with important functions in plant stress adaptation. Exploiting the functional genomics and bioinformatics studies paved the way in understanding the relationship between genotype and phenotype of an organism suffering from environmental stress. Future research programs can be focused on the development of transgenic plants with enhanced stress tolerance in field conditions based upon the outcome of genomic approaches and knowing the mystery of nucleotides sequences hidden in cells.

The extreme environmental conditions disturbs plant growth and development, restricting the genetic potential and yield losses in plants. It is necessary to understand plant responses to stresses that disturbs the homeostatic equilibrium. Newer genes and markers need to be identified to make heat stress tolerant crops and comparative studies in ancestral plant groups can help to understand the complex biological processes. The plants cope with adverse temperature conditions by regulating various mechanism and sustains high temperature stress by producing heat shock proteins (HSPs) and innate immunity and abiotic stress tolerance in higher plants. In this context, in silico methods were used to discover new EST derived microsatellite markers associated with heat stress tolerance in Solanummelongenafor plant breeding programme as they are directly linked to functional genes. The publically available EST collection of S. melongena were explored to identify EST linked markers associated with heat tolerance. Approximately 529 EST sequences having repeats were analyzed for stress tolerance using insilico methods. These were categorized into 11 different protein superfamilies implicated in stress tolerance based on conserved domain and protein functions. Among these, five unique EST sequences pertains to HSP90 superfamily associated with heat stress tolerance were found having a microsatellite marker. These are potential markers useful for marker assisted selection in breeding programs and also facilitate the targeted research for development of heat stress tolerant crops.

Pigeonpea (Cajanus cajan(L) Millsp.) is an important food legume crop of rain fed agriculture in the arid and semiarid tropics of the world. It has deep and extensive root system which serves a number of important physiological and metabolic functions in plant development and growth. In order to identify genes associated with pigeonpea root, ESTs were generated from the root tissues of pigeonpea (GRG-295 genotype) by normalized cDNA library. A total of 105 high quality ESTs were generated by sequencing of 250 random clones which resulted in 72 unigenes comprising 25 contigs and 47 singlets. The ESTs were assigned to 9 functional categories on the basis of their putative function. In order to validate the possible expression of transcripts, four genes, namely,S-adenosylmethionine synthetase,phosphoglycerate kinase,serine carboxypeptidase, andmethionine aminopeptidase, were further analyzed by reverse transcriptase PCR. The possible role of the identified transcripts and their functio...

Plants in nature may face a wide range of favorable or unfavorable biotic and abiotic factors during their life cycle. Any of these factors may cause stress in plants; therefore, they have to be more adaptable to stressful environments and must acquire greater response to different stresses. The objective of this study is to retrieve and arrange data from the literature in a standardized electronic format for the development of information resources on potential stress responsive genes in Arabidopsis thaliana. This provides a powerful mean for manipulation, comparison, search, and retrieval of records describing the nature of various stress responsive genes in Arabidopsis thaliana. The database is based exclusively on published stress tolerance genes associated with plants.

Ploidy-variable species allow direct inference of the effects of chromosome copy number on fundamental evolutionary processes. While an abundance of theoretical work suggests polyploidy should leave distinct population genomic signatures, empirical data remains sparse. We sequenced ~300 individuals from 39 populations of Arabidopsis arenosa, a naturally diploid-autotetraploid species. We find the impacts of polyploidy on population genomic processes are subtle yet pervasive, including reduced efficiency of purifying selection, differences in linked selection, and rampant gene flow from diploids. Initial masking of deleterious mutations, faster rates of nucleotide substitution, and interploidy introgression likely conspire to shape the evolutionary potential of polyploids.

While it is possible to interpret microarray experiments a single gene at a time, most studies generate long lists of differentially expressed genes whose interpretation requires the integration of prior biological knowledge. This prior knowledge is stored in various public and private databases and covers several aspects of gene function and biological information. In this review, we will describe the tools and places where to find prior accurate biological information and how to process and incorporate them to interpret microarray data analyses. Here, we highlight selected tools and resources for gene class level ontology analysis (Section 2), gene coexpression analysis (Section 3), gene network analysis (Section 4), biological pathway analysis (Section 5), analysis of transcriptional regulation (Section 6), and omics data integration (Section 7). The overall goal of this review is to provide researchers with tools and information to facilitate the interpretation of microarray data.

Low temperature (LT) in spring has become one of the principal abiotic stresses that restrict the growth and development of wheat.  Diverse analyses were performed to investigate the mechanism underlying the response of wheat grain development to LT stress during booting.  These included morphological observation, measurements of starch synthase activity, and determination of amylose and amylopectin content of wheat grain after exposure to treatment with LT during booting.  Additionally, proteomic analysis was performed using tandem mass tags (TMT).  Results showed that the plumpness of wheat grains decreased after LT stress.  Moreover, the activities of sucrose synthase (SuS, EC 2.4.1.13) and ADP-glucose pyrophosphorylase (AGPase, EC 2.7.7.27) exhibited a significant reduction, leading to a significant reduction in the contents of amylose and amylopectin.  A total of 509 differentially expressed proteins (DEPs) were identified by proteomics analysis.  The Gene Ontology (GO) enrichment analysis showed that the protein difference multiple in the nutritional repository activity was the largest among the molecular functions, and the up-regulated seed storage protein (SSP) played an active role in the response of grains to LT stress and subsequent damage.  The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that LT stress reduced the expression of DEPs such as sucrose phosphate synthase (SPS), glucose-1-phosphate adenylyltransferase (glgC), and β-fructofuranosidase (FFase) in sucrose and starch metabolic pathways, thus affecting the synthesis of grain starch.  In addition, many heat shock proteins (HSPs) were found in the protein processing in endoplasmic reticulum pathways, which can resist some damage caused by LT stress.  These findings provide a new theoretical foundation for elucidating the underlying mechanism governing wheat yield development after exposure to LT stress in spring.

Plant breeders and geneticists use molecular marker-assisted selection also called as MAS as a useful approach for breeding of plant to make selection more efficient and speed up the breeding cycle. MAS can be more efficient, effective, and reliable than phenotypic selection. Molecular markers are useful to identify the economically important traits in the breeding population for further manipulation in a short time. Due to the applicability of markers at the seedling stage ensuring high precision at the reduced level of cost, marker-assisted selection offer the chances to improve responses from selection. The MAS using DNA level polymorphism accelerate the pace of selection. The main marker technologies applied are chiefly co-dominant markers i.e. microsatellite markers/SSR (Simple Sequence Repeats) marker, RFLP (Restriction Fragment Length Polymorphism) marker and SNPs (Single nucleotide polymorphisms). This review overviews the various MAS technologies and their applications in crop improvement programs.

Glycogenes regulate a wide array of biological processes in the development of organisms as well as different diseases such as cancer, primary open-angle glaucoma, and renal dysfunction. The objective of this study was to explore the role of differentially expressed glycogenes (DEGGs) in three major tissues such as brain, muscle, and liver using mouse RNA-seq data, and we identified 579, 501, and 442 DEGGs for brain versus liver (BvL579), brain versus muscle (BvM501), and liver versus muscle (LvM442) groups. DAVID functional analysis suggested inflammatory response, glycosaminoglycan metabolic process, and protein maturation as the enriched biological processes in BvL579, BvM501, and LvM442, respectively. These DEGGs were then used to construct three interaction networks by using GeneMANIA, from which we detected potential hub genes such asPEMTandHPXN(BvL579),IGF2andNID2(BvM501), andSTAT6andFLT1(LvM442), having the highest degree. Additionally, our community analysis results suggest...

Low transformation efficiency is one of the main limiting factors in the establishment of genetic transformation of wheat via Agrobacterium tumefaciens. To determine more favorable conditions for T-DNA delivery and explant regeneration after infection, this study investigated combinations of acetosyringone concentration and pH variation in the inoculation and co-cultivation media and co-culture temperatures using immature embryos from two Brazilian genotypes (BR 18 Terena and PF 020037). Based on transient expression of uidA, the most favorable conditions for T-DNA delivery were culture media with pH 5.0 and 5.4 combined with co-culture temperatures of 22 °C and 25 °C, and a 400 mM acetosyringone supplement. These conditions resulted in blue foci in 81% of the embryos. Media with more acidic pH also presented reduced A. tumefaciens overgrowth during co-culture, and improved regeneration frequency of the inoculated explants. BR 18 Terena was more susceptible to infection by A. tumefaciens than PF 020037. We found that it is possible to improve T-DNA delivery and explant regeneration by adjusting factors involved in the early stages of A. tumefaciens infection. This can contribute to establishing a stable transformation procedure in the future.

Caffeic acid o-methyltransferase (COMT) is one of the important enzymes controlling lignin monomer production in plant cell wall synthesis. Analysis of the genome sequence of the new grass model Brachypodium distachyon identified four COMT gene homologs, designated as BdCOMT1, BdCOMT2, BdCOMT3, and BdCOMT4. Phylogenetic analysis suggested that they belong to the COMT gene family, whereas syntenic analysis through comparisons with rice and sorghum revealed that BdCOMT4 on Chromosome 3 is the orthologous copy of the COMT genes well characterized in other grass species. The other three COMT genes are unique to Brachypodium since orthologous copies are not found in the collinear regions of rice and sorghum genomes. Expression studies indicated that all four Brachypodium COMT genes are transcribed but with distinct patterns of tissue specificity. Full-length cDNAs were cloned in frame into the pQE-T7 expression vector for the purification of recombinant Brachypodium COMT proteins. Bioche...

Unlike animals, plants cannot flee, fight or hide from predators. Plants lack mobile defender cells or an adaptive immune system and have instead evolved defences based on pre-formed barriers and inducible cellular responses regulated by local and systemic signals. The interaction between pathogen effectors and these defences sets up an intriguing molecular arms race between plants and pathogenic fungi, bacteria, viruses, viroids and nematodes. Plant surfaces present formidable physical and chemical barriers as the first line of defences against pathogens. Waxy cuticles, cellulosic or lignified cell walls and antimicrobial phytoanticipins and defensins exclude most microbes. In a classic experiment, JC Walker and colleagues showed that brown onions resist smudge disease, caused by the fungus Colletotrichum circinans, because of the antifungal phenolic protocatechuic acid in their dry outer leaves 1 . White onions, lacking these compounds, are susceptible. Following damage or infection, plant metabolites present in inactive forms may be converted to more inhibitory substances, including glucosinolates, cyanides and toxic quinones 2 . Avenacins, triterpenoid saponins found in the outer cortex of oat roots, confer resistance to take-all caused by non-specific races of Gauemannomyces graminis 3 . The virulent oat-specific pathogen G. graminis var. avenae produces the detoxifying enzyme avenacinase, and races of the pathogen from other cereals acquired virulence on oats when transformed with the G. graminis var. avenae avenacinase gene 4 , while oat mutants defective in avenacin biosynthesis became susceptible 5 . Plant surfaces are also equipped with arrays of cell surface pattern recognition receptors (PRRs) that detect pathogen-associated molecular patterns (PAMPs). Recognition activates mitogenactivated protein kinase (MAPK) cascades that both positively and negatively regulate PAMP-triggered immunity (PTI) within the cell . PTI includes responses that disrupt parasitism, activate defensive cell death programs, defence-related gene expression and antibiotic accumulation, and reinforce plant cell walls 7,9 .

During the period 2002-10, FAO undertook an intense activity of biosafety capacity development, largely centred on enhancing the capacities of regulators and other technical staff involved in the implementation of biosafety frameworks, along with other components. The training programme was tailored to meet the needs of a very specific audience: biosafety regulators, policy-makers and members of national biosafety committees, with diverse educational backgrounds, not necessarily well versed in all the biosafety-related fields. The training courses therefore aimed to: i) offer background knowledge critical in the process of reviewing biosafety dossiers and biosafety-related decision-making ; ii) provide acquaintance with concepts and methodologies relevant to risk analysis of genetically modified organism (GMO) release and biosafety management. The training programme consisted of the following modules: Module A: Introduction to molecular biology and genetic engineering. This reviews ...

Wintersweet (Chimonanthus praecox) is familiar as a garden plant and woody ornamental flower. On account of its unique flowering time and strong fragrance, it has a high ornamental and economic value. Despite a long history of human cultivation, our understanding of wintersweet genetics and molecular biology remains scant, reflecting a lack of basic genomic and transcriptomic data. In this study, we assembled three cDNA libraries, from three successive stages in flower development, designated as the flower bud with displayed petal, open flower and senescing flower stages. Using the Illumina RNA-Seq method, we obtained 412,928,950,404,912,954 qualified Illumina reads, respectively, for the three successive stages. The pooled reads from all three libraries were then assembled into 106,995 transcripts, 51,793 of which were annotated in the NCBI non-redundant protein database. Of these annotated sequences, 32,649 and 21,893 transcripts were assigned to gene ontology categories and clusters of orthologous groups, respectively. We could map 15,587 transcripts onto 312 pathways using the Kyoto Encyclopedia of Genes and Genomes pathway database. Based on these transcriptomic data, we obtained a large number of candidate genes that were differentially expressed at the open flower and senescing flower stages. An analysis of differentially expressed genes involved in plant hormone signal transduction pathways indicated that although flower opening and senescence may be independent of the ethylene signaling pathway in wintersweet, salicylic acid may be involved in the regulation of flower senescence. We also succeeded in isolating key genes of floral scent biosynthesis and proposed a biosynthetic pathway for monoterpenes and sesquiterpenes in wintersweet flowers, based on the annotated sequences. This comprehensive transcriptomic analysis presents fundamental information on the genes and pathways which are involved in flower development in wintersweet. And our data provided a useful database for further research of wintersweet and other Calycanthaceae family plants.

Plant mutagenesis is rapidly coming of age in the aftermath of recent developments in high-resolution molecular and biochemical techniques. By combining the high variation of mutagenised populations with novel screening methods, traits that are almost impossible to identify by conventional breeding are now being developed and characterised at the molecular level. This paper provides a comprehensive overview of the various techniques and workflows available to researchers today in the field of molecular breeding, and how these tools complement the ones already used in traditional breeding. Both genetic (Targeting Induced Local Lesions in Genomes; TILLING) and phenotypic screens are evaluated. Finally, different ways of bridging the gap between genotype and phenotype are discussed.

Cowpea is an important tropical crop. It provides a large proportion of the food resource for the African human population and their livestock. The yield and quality of cowpea have been dramatically improved through traditional breeding strategies for the past few decades. However, reports of heritability estimates for early growth of cowpea are rare. We designed a simple experiment to estimate the broad-sense heritability of early growth. We randomly selected 15 cowpea varieties among a total of 5000 cowpea accessions maintained in the cowpea breeding facility at the University of California, Riverside to examine the genetic determination of early growth of cowpea (measured as the height at day five after seeding). The estimated broad-sense heritability on the individual plant basis is 0.2190. However, the corresponding estimate on the plant mean basis (average of four plants) is 0.5198, which is very high for a quantitative trait. The high heritability may explain why traditional ...

Depuis une vingtaine d'années, les nombreuses recherches focalisées sur la plante modèle Arabidopsis thaliana ont permis des progrès considérables dans notre connaissance des bases moléculaires de la biologie des plantes. Avec l'achèvement du séquençage du génome d'Arabidopsis [1], la palette complète des outils de génétique et de génomique maintenant disponibles devrait encore accélérer le rythme des découvertes. De son côté, le riz fait l'objet de rapides développements génomiques en tant que modèle pour les monocotylédones. Cependant, il est clair que ces deux espèces ne suffisent pas pour représenter le monde végétal dans toute sa diversité biologique [2]. Ce fait est particulièrement bien illustré par le cas des Légumineuses (Fabacées) qui représentent l'un des taxons végétaux les plus importants, tant du point de vue de la biologie et de l'écologie fondamentales que du point de vue agronomique et environnemental. Summary : The legume species Medicago truncatula is now a model plant used worldwide , and in particular for molecular genetic studies of the endomycorrhizal and nitrogen-fixing root symbioses, associations which do not occur with Arabidopsis. Large-scale projects for M. truncatula genomics have been initiated within the international community and essential tools are being currently developed for structural genomics (genome mapping, BAC libraries, genome sequencing) and functional genomics (ESTs, microarrays, mutant collections), along with the development of bioinformatics resources. Comparative genomics studies suggest a relatively high level of synteny between legume genomes. It can therefore be anticipated that M. truncatula will be used as a nodal legume species to help in the identification of agronomically important genes in crop legumes, e.g. genes for root symbioses, disease/pest resistance, plant architecture, seed quality, and production of specific secondary metabolites.

Single nucleotide polymorphisms (SNPs) are the most common DNA polymorphisms in plant and animal genomes. SNPs were identified in the allele sequences of up to 12 sunflower (Helianthus annuus L.) genotypes for a genomewide sample of 81 loci originally mapped using restriction fragment length polymorphism (RFLP) markers. The RFLP loci anchor a dense RFLP linkage map and second-generation linkage maps constructed using simple sequence repeats and other high-throughput DNA markers. The goal of this study was to develop high-throughput SNP markers for the cDNA-RFLP loci for linkage mapping, diversity analysis, and molecular breeding and genomics research. The specific objectives were to: (i) develop FP-TDI or fluorescent primer-extension SNP assays for the cDNA-RFLP loci; (ii) validate the SNP markers by screening 12 sequenced (reference) genotypes; and (iii) assess the utility and polymorphism rate of the SNP markers across a genetically diverse panel of unsequenced wild and domesticated sunflower genotypes. I am grateful to my family, Mike, Maggie, and Katie Freeman, for their love and support during my time at Oregon State University. Without them I could not have accomplished as much as I have in life. My partner in crime and life, Adam Barrios, will be forever appreciated for his unwavering support during my graduate career. Thank you for continually supporting me in my endeavors. CONTRIBUTION OF AUTHORS Dr. Steven Knapp proposed and supervised this research. Judith Kolkman obtained the preliminary data, including sequence alignments, from which this research was built on. Loren Rieseberg and Jerry Miller generously supplied seed samples and DNA samples were supplied by Advanta Semillas.

A total of 16 species of Garcinia have been investigated in the 50-ha permanent ecological plot in Pasoh Forest Reserve, Negeri Sembilan. From a total of 4,035 individual stems of Garcinia recorded, the most abundant species is G. malaccensis (902 stems) while G. parvifolia (1.91 m 2 of basal area) is the most dominant species. This study has shown a trend in which species diversity is strongly influenced by variation of the soil types. More than half (54.7%) of the trees and seedlings are found on the welldrained soil developed from alluvial parent material. While the poorly-drained soil of the acidic and ferric substrates are the least preferred. The changes in population size in the period of ten years were relatively low at 0.78% with an annual mortality rate at 1.90% per year of which 85.8% of the dead trees came from 1.0-5.0 cm dbh trees. A total of 439 new individuals were recorded and 397 individuals were recruited alone in 1990-1995 period. The mean dbh increment was low at 8 mm per year for all sub-class dbh but adult trees grew at 12.2 mm per year. The low mortality rate indicates no major disturbance in this virgin forest and slow growth increment was a main character in tropical primary forest.

MicroRNAs (miRNAs) are endogenous non-coding RNAs that play important regulatory roles in animals and plants by targeting mRNAs for cleavage or translational repression. Small RNAs are classified into different types by their biogenesis and mode of action, such as miRNAs, siRNAs, piRNAs, and snoRNAs. In the case of miRNAs, this specific type regulates gene expression in plants and animals by targeting mRNAs for cleavage and translational repression, respectively. Diverse miRNAs regulate plant development, metabolism, and responses to biotic and abiotic stresses. The identification of miRNAs has been accomplished in diverse species, organs and developmental or diverse biotic and abiotic stress conditions. Novel massive sequencing techniques and further bioinformatics analysis have allowed the identification of hundreds of miRNAs in Arabidopsis thaliana, Oryza sativa, Malus domestica, Zea mays, Solanum lycopersicum, and other plants. Functional characterization of a given miRNA in a specific biological context has shown their role in the finetuning mechanisms of posttranscriptional gene regulation. In this chapter, besides making a summary of genome-wide miRNA profiling in plants, we describe how gain and loss of function approaches influence plant phenotypes that affect development, physiology or stress responses, pointing to miRNAs as effective tools for the generation of new plant phenotypes that improve plant productivity and conservation.

Microsatellites or SSRs (simple sequence repeats) are ubiquitous short tandem duplications occurring in eukaryotic organisms. These sequences are among the best marker technologies applied in plant genetics and breeding. The abundant genomic, BAC, and EST sequences available in databases allow the survey regarding presence and location of SSR loci. Additional information concerning primer sequences is also the target of plant geneticists and breeders. In this paper, we describe a utility that integrates SSR searches, frequency of occurrence of motifs and arrangements, primer design, and PCR simulation against other databases. This simulation allows the performance of global alignments and identity and homology searches between different amplified sequences, that is, amplicons. In order to validate the tool functions, SSR discovery searches were performed in a database containing 28 469 nonredundant rice cDNA sequences.

Bursera comprises ~100 tropical shrub and tree species, with the center of the species diversification in Mexico. The genomic resources developed for the genus are scarce, and this has limited the study of the gene flow, local adaptation, and hybridization dynamics. In this study, based on ~155 million Illumina paired-end reads per species, we performed a de novo genome assembly and annotation of three Bursera species of the Bullockia section: Bursera bipinnata, Bursera cuneata, and Bursera palmeri. The total lengths of the genome assemblies were 253, 237, and 229 Mb for B. cuneata, B. palmeri, and B. bipinnata, respectively. The assembly of B. palmeri retrieved the most complete and single-copy BUSCOs (87.3%) relative to B. cuneata (86.5%) and B. bipinnata (76.6%). The ab initio gene prediction recognized between 21,000 and 32,000 protein-coding genes. Other genomic features, such as simple sequence repeats (SSRs), were also detected. Using the de novo genome assemblies as a refere...

The benzylisoquinoline alkaloid papaverine, synthesized in low amount in most of the opium poppy varieties of Papaver somniferum, is used as a vasodilator muscle relaxant and antispasmodic. Papaverine biosynthesis remains controversial as two different routes utilizing either (S)-coclaurine or (S)-reticuline have been proposed with uncharacterized intermediate steps. In an attempt to elucidate papaverine biosynthesis and identify putative genes involved in uncharacterized steps, we carried out comparative transcriptome analysis of high papaverine mutant (pap1) and normal cultivar (BR086) of P. somniferum. This natural mutant synthesizes more than 12-fold papaverine in comparison to BR086. We established more than 238 Mb transcriptome data separately for pap1 and BR086. Assembly of reads generated 127,342 and 106,128 unigenes in pap1 and BR086, respectively. Digital gene expression analysis of transcriptomes revealed 3,336 differentially expressing unigenes. Enhanced expression of (S)-norcoclaurine-6-O-methyltransferase (6OMT), (S)-39-hydroxy-N-methylcoclaurine 49-O-methyltransferase (49OMT), norreticuline 7-O-methyltransferase (N7OMT) and down-regulation of reticuline 7-O-methyltransferase (7OMT) in pap1 in comparison to BR086 suggest (S)-coclaurine as the route for papaverine biosynthesis. We also identified several methyltransferases and dehydrogenases with enhanced expression in pap1 in comparison to BR086. Our analysis using natural mutant, pap1, concludes that (S)-coclaurine is the branch-point intermediate and preferred route for papaverine biosynthesis. Differentially expressing methyltransferases and dehydrogenases identified in this study will help in elucidating complete biosynthetic pathway of papaverine. The information generated will be helpful in developing strategies for enhanced biosynthesis of papaverine through biotechnological approaches.

Virus-induced gene silencing (VIGS) is used to down-regulate endogenous plant genes. VIGS efficiency depends on viral proliferation and systemic movement throughout the plant. Although tobacco rattle virus (TRV)-based VIGS has been successfully used in petunia (Petunia × hybrida), the protocol has not been thoroughly optimized for efficient and uniform gene down-regulation in this species. Therefore, we evaluated six parameters that improved VIGS in petunia. Inoculation of mechanically wounded shoot apical meristems induced the most effective and consistent silencing compared to other methods of inoculation. From an evaluation of ten cultivars, a compact petunia variety, 'Picobella Blue', exhibited a 1.8-fold higher CHS silencing efficiency in corollas. We determined that 20°C day/18°C night temperatures induced stronger gene silencing than 23°C/18°C or 26°C/ 18°C. The development of silencing was more pronounced in plants that were inoculated at 3-4 versus 5 weeks after sowing. While petunias inoculated with pTRV2-NbPDS or pTRV2-PhCHS showed very minimal viral symptoms, plants inoculated with the pTRV2 empty vector (often used as a control) were stunted and developed severe necrosis, which often led to plant death. Viral symptoms were eliminated by developing a control construct containing a fragment of the green fluorescent protein (pTRV2-sGFP). These optimization steps increased the area of chalcone synthase (CHS) silencing by 69 % and phytoene desaturase (PDS) silencing by 28 %. This improved VIGS protocol, including the use of the pTRV2-sGFP control plants, provides stronger down-regulation for high-throughput analyses of gene function in petunia.

Salinity stress limits crop yield affecting plant growth and restricting the use of land. As world population is increasing at alarming rate, agricultural land is shrinking due to industrialization and/or habitat use. Hence, there is a need to utilize salt affected land to meet the food requirement. Although some success has been achieved through conventional breeding, but its use is limited due to reproductive barrier and scarcity of genetic variations among major crops. The genetic engineering has proven a revolutionary technique to generate salt tolerant plants as one can transfer desired gene from any genetic resource and/or alter the expression of existing gene(s). There are examples of improved salinity tolerance in various crop plants through the use of genetic engineering. However, there is a further need of improvement for successful release of salt tolerant cultivars at field level. In this research, we have given a detailed update on production of salttolerant plants through genetic engineering. Future prospects and concerns, along with the importance of novel techniques, as well as plant breeding are also discussed.

Salinity stress limits crop yield affecting plant growth and restricting the use of land. As world population is increasing at alarming rate, agricultural land is shrinking due to industrialization and/or habitat use. Hence, there is a need to utilize salt affected land to meet the food requirement. Although some success has been achieved through conventional breeding, but its use is limited due to reproductive barrier and scarcity of genetic variations among major crops. The genetic engineering has proven a revolutionary technique to generate salt tolerant plants as one can transfer desired gene from any genetic resource and/or alter the expression of existing gene(s). There are examples of improved salinity tolerance in various crop plants through the use of genetic engineering. However, there is a further need of improvement for successful release of salt tolerant cultivars at field level. In this research, we have given a detailed update on production of salttolerant plants thro...

Protein-protein interactions (PPIs) and multiprotein complexes perform central roles in the cellular systems of all living organisms. In humans, disruptions of the normal patterns of PPIs and protein complexes can be causative or indicative of a disease state. Recent developments in the biological applications of mass spectrometry (MS)-based proteomics have expanded the horizon for the application of systematic large-scale mapping of physical interactions to probe disease mechanisms. In this review, we examine the application of MS-based approaches for the experimental analysis of PPI networks and protein complexes, focusing on the different model systems (including human cells) used to study the molecular basis of common diseases such as cancer, cardiomyopathies, diabetes, microbial infections, and genetic and neurodegenerative disorders. Index Terms PPI, Complex diseases, Networks

With the ever-increasing dilemma experienced by the agricultural sector, due to the ever-increasing and continuous demand for food, feed, and fiber amidst population growth and declining conditions that favor agricultural productivity, there is a need to develop means of producing agricultural products more sustainably through highly efficient and effective models. Digital technologies in the agriculture sector, present a ray of hope with tools such as Geographical Information System (GIS), Remote Sensing (RS), Artificial Intelligence (AI), Precision Agriculture (PA), etc. The incorporation of digital technologies in agriculture may help solve the problem of global warming and the ever-increasing population associated with declining levels of natural resources. Hence, digital agriculture has the potential to revolutionize sustainable crop production substantially, to achieve the goal of 'zero hunger'. Therefore, the present review highlights some of the digital technology tools and their role in sustainable crop production if utilized efficiently.

Halophytes are plants which naturally survive in saline environment. They account for ∼1% of the total flora of the world. They include both dicots and monocots and are distributed mainly in arid, semi-arid inlands and saline wet lands along the tropical and subtropical coasts. Salinity tolerance in halophytes depends on a set of ecological and physiological characteristics that allow them to grow and flourish in high saline conditions. The ability of halophytes to tolerate high salt is determined by the effective coordination between various physiological processes, metabolic pathways and protein or gene networks responsible for delivering salinity tolerance. The salinity responsive proteins belong to diverse functional classes such as photosynthesis, redox homeostasis; stress/defense, carbohydrate and energy metabolism, protein metabolism, signal transduction and membrane transport. The important metabolites which are involved in salt tolerance of halophytes are proline and proline analog (4-hydroxy-N-methyl proline), glycine betaine, pinitol, myo-inositol, mannitol, sorbitol, O-methylmucoinositol, and polyamines. In halophytes, the synthesis of specific proteins and osmotically active metabolites control ion and water flux and support scavenging of oxygen radicals under salt stress condition. The present review summarizes the salt tolerance mechanisms of halophytes by elucidating the recent studies that have focused on proteomic, metabolomic, and ionomic aspects of various halophytes in response to salinity. By integrating the information from halophytes and its comparison with glycophytes could give an overview of salt tolerance mechanisms in halophytes, thus laying down the pavement for development of salt tolerant crop plants through genetic modification and effective breeding strategies.

While it is possible to interpret microarray experiments a single gene at a time, most studies generate long lists of differentially expressed genes whose interpretation requires the integration of prior biological knowledge. This prior knowledge is stored in various public and private databases and covers several aspects of gene function and biological information. In this review, we will describe the tools and places where to find prior accurate biological information and how to process and incorporate them to interpret microarray data analyses. Here, we highlight selected tools and resources for gene class level ontology analysis (Section 2), gene coexpression analysis (Section 3), gene network analysis (Section 4), biological pathway analysis (Section 5), analysis of transcriptional regulation (Section 6), and omics data integration (Section 7). The overall goal of this review is to provide researchers with tools and information to facilitate the interpretation of microarray data.

Lettuce is a major vegetable in western countries. Mutations generated genetic variations and played an important role in the domestication of the crop. Many traits derived from natural and induced mutations, such as dwarfing, early flowering, male sterility, and chlorophyll deficiency, are useful in physiological and genetic studies. Mutants were also used to develop new lettuce products including miniature and herbicide-tolerant cultivars. Mutant analysis was critical in lettuce genomic studies including identification and cloning of disease-resistance genes. Mutagenesis combined with genomic technology may provide powerful tools for the discovery of novel gene alleles. In addition to radiation and chemical mutagens, unconventional approaches such as tissue or protoplast culture, transposable elements, and space flights have been utilized to generate mutants in lettuce. Since mutation breeding is considered nontransgenic, it is more acceptable to consumers and will be explored mor...

De novo assemblies do not have the possibility of quality control with an external sequence. In fact, accuracy and reliability of these assemblies is highly affected by sequencing errors and mis-assemblies. Here, a frequency- based algorithm is developed in Ruby and intended to discern assembly errors from polymorphisms/read errors and then edit or remove the misassembled read(s) to provide more but highly reliable contigs. The software reads and writes the ACE assembly format. Transcriptome and genome assemblies were tested.

The Generation Challenge programme (GCP) is a global crop research consortium directed toward crop improvement through the application of comparative biology and genetic resources characterization to plant breeding. A key consortium research activity is the development of a GCP crop bioinformatics platform to support GCP research. This platform includes the following: (i) shared, public platform-independent domain models, ontology, and data formats to enable interoperability of data and analysis flows within the platform; (ii) web service and registry technologies to identify, share, and integrate information across diverse, globally dispersed data sources, as well as to access high-performance computational (HPC) facilities for computationally intensive, high-throughput analyses of project data; (iii) platform-specific middleware reference implementations of the domain model integrating a suite of public (largely open-access/-source) databases and software tools into a workbench to...

Multigeneic QTL present significant problems to analysis. Resistance to soybean (Glycine max (L) Merr.) sudden death syndrome (SDS) caused by Fusarium virguliforme was partly underlain by QRfs2 that was clustered with, or pleiotropic to, the multigeneic rhg1 locus providing resistance to soybean cyst nematode (SCN; Heterodera glycines). A group of five genes were found between the two markers that delimited the Rfs2/rhg1 locus. One of the five genes was predicted to encode an unusual diphenol oxidase (laccase; EC 1.10.3.2). The aim of this study was to characterize this member of the soybean laccase gene-family and explore its involvement in SDS resistance. A genomic clone and a full length cDNA was isolated from resistant cultivar 'Forrest' that were different among susceptible cultivars 'Asgrow 3244' and 'Williams 82' at four residues R/H168, I/M271, R/H330, E/K470. Additional differences were found in six of the seven introns and the promoter region. Transcript abundance (TA) among genotypes that varied for resistance to SDS or SCN did not differ significantly. Therefore the protein activity was inferred to underlie resistance. Protein expressed in yeast pYES2/NTB had weak enzyme activity with common substrates but good activity with root phenolics. The Forrest isoform may underlie both QRfs2 and rhg1.

ÖZET : Bitkiler doğada canlılıklarını sürdürebilmek için çeşitli savunma mekanizmaları geliştirmişlerdir. RNA ortamlı antiviral savunma mekanizması olarak bilinen Virüs Kaynaklı Gen Susturma (VIGS) bu mekanizmalardan biridir. Virüsle enfekte olmuş bitkilerde, virüs genomuna karşı savunma mekanizması çalışmaktadır. Aynı zamanda hedef geni taşıyan virüs vektörleri homolog içgen mRNA'larını da bozabilmektedir. Bununla birlikte VIGS genin fonksiyonunu anlamada ve fonksiyonel genomik analizleri için kullanılmaktadır. Birçok bitki için virüs vektörleri geliştirilmiştir. Son zamanlarda tahıllar için Arpa Çizgili Mozaik Virüsü (Barley Stripe Mosaic Virus = BSMV) kullanılmaya başlanmıştır. BSMV geniş bir konukçu aralığına sahip pozitif polariteli bir RNA virüsü olup, Hordeivirüs genusunun bir üyesidir. Üç farklı genom RNA (α, β ve γ)' dan oluşmaktadır. Bu çalışmada BSMV vektörünün çalışma prensibi ve bazı tahıllarda BSMV-VIGS'in uygulamaları hakkında bilgi verilmiştir.

Sabev, P., Valkova, N. & Todorovska, E. G. (2020). Molecular markers and their application in cotton breeding: progress and future perspectives. Bulg. J. Agric. Sci., 26 (4), 816–828 Cotton is the most important natural fiber plant worldwide and a valued cash crop. It is also a significant source of oil and protein. The conventional breeding for genetic improvement has its limitations due mostly to the lack of knowledge about polygenic traits such as yield productivity, fiber quality, resistance to diseases, and adverse growing conditions. The use of molecular markers is one of the significant developments in the field of genetics, allowing faster detection and exploitation of DNA polymorphisms. Recent developments, including the sequencing of the genome of G. raimondii, G. arboreum, G. hirsutum, and G. barbadense open new possibilities for selection of the best traits present in the genome of cotton. This article gives an overview of various DNA markers used in cotton. They play a ...

The promise of association genetics to identify genes or genomic regions controlling complex traits has generated a flurry of interest. Such phenotype-genotype associations could be useful to accelerate tree breeding cycles, increase precision and selection intensity for late expressing, low heritability traits. However, the prospects of association genetics in highly heterozygous undomesticated forest trees can be severely impacted by the presence of cryptic population and pedigree structure. To investigate how to better account for this, we compared the GLM and five combinations of the Unified Mixed Model (UMM) on data of a low-density genome-wide association study for growth and wood property traits carried out in a Eucalyptus globulus population (n = 303) with 7,680 Diversity Array Technology (DArT) markers. Model comparisons were based on the degree of deviation from the uniform distribution and estimates of the mean square differences between the observed and expected p-values of all significant marker-trait associations detected. Our analysis revealed the presence of population and family structure. There was not a single best model for all traits. Striking differences in detection power and accuracy were observed among the different models especially when population structure was not accounted for. The UMM method was the best and produced superior results when compared to GLM for all traits. Following stringent correction for false discoveries, 18 marker-trait associations were detected, 16 for tree diameter growth and two for lignin monomer composition (S:G ratio), a key wood property trait. The two DArT markers associated with S:G ratio on chromosome 10, physically map within 1 Mbp of the ferulate 5-hydroxylase (F5H) gene, providing a putative independent validation of this marker-trait association. This study details the merit of collectively integrate population structure and relatedness in association analyses in undomesticated, highly heterozygous forest trees, and provides additional insights into the nature of complex quantitative traits in Eucalyptus.

The objective of this work was to determine the preference and the demographic parameters of cowpea aphid ( Aphis craccivora ) in advanced lines of semiprostrate cowpea ( Vigna unguiculata ), and to verify both the existance and type of plant resistance to this insect attack. Aphid preference to 20 cowpea genotypes was tested in a randomized complete block design, with six replicates. Lines showing the highest levels of resistance and susceptibility in the preference tests were selected for use in the demographic parameter tests. Based on aphid birth rate and mortality, fertility life tables were constructed. The MNCO4-792F-123 line was classified as resistant. The lowest values for cowpea aphid intrinsic and finite growth rates were obtained for the MNCO4-792F-123, and the highest ones for MNCO4-782F-108 and 'BRS Novaera' genotypes, whose levels exceeded those of 'Vita 7', the standard genotype for susceptibility. The MNCO4-792F-123 line is the least favorable for t...