Cloning Of Rice Dna And Identification Of Transfer-Rna Gene Clones

THE rapid development of recombinant DNA technology has brought forth a revolution in biology'>", it aids us to have a closer look at the 'way genes are organized, eS11 ecially in the complex eucaryotic genornes'<", Although many animal and yeast genes have been studied in detail using recombinant DNA technology, plant genes have seldom been targets for such studie., Germination is an ideal process to study gene expression .because it effects a . shift in the metabolic status of seeds from a state of 'dormancy to an active one. AJ;l understanding of gene organization and regulation darin.g germination can be accomplblted by molecular cloning of DNA from seeds lik.e rice. To study the status of histone, rRNA tRNA and other genes in the rice genome, a general method was developed to clone eucarvotic DNA in a' plasmid vector pBR 322. This essentially ~ involves the following steps. The rice embryo and plasmid pBR 322 DNAs were cut witll restriction endonuclease Bam Hi to generate stick.Y ends, The plasmid DNA was puosphatased, the DNA~ ware a~·tnealed and joined 'by T4 phage DNA ligase. The recombinant DNA molecules thus produced were transjerred into E. coli and colonies containing them Were selected by their sensitivity to tetracycline and resistance to ampicillin, Two clones were identified . 2S haVing tRNA genes by hybridization of the DNA in the clones \vitl1 32P-la.belled rice tRNAs.

Podiatry scope of practice: The role of gender in identification as a specialist

Background Ensuring efficient and effective delivery of health care to an aging population has been a major driver for a review of the health workforce in Australia. As part of this review a new National Registration and Accreditation Scheme (NRAS) has evolved with one goal being to improve workforce flexibility. With increased flexibility there have been discussions about the role specialist scopes of practice plays. This study explored the role of gender and other work related characteristics in relation to contemporary scope of podiatry practice and specialisation in Australia. Methods A cross sectional survey was administered through an on-line survey tool on behalf of the Australasian Podiatry Council. Descriptive data was collected over a three-week period. Queensland University of Technology Human Research Ethics approval was sought and confirmed exemption from review, exemption number 1400000791. Results Of the podiatrists participating in this survey (n=218), they were predominately female (66%), early career (34%, 0-9 years) and work in private practices (78%) in multi-podiatrists centres (41%). Relationship between clinical activities performed and “self-perception” of performing a “specialist role” was significant for practitioners who undertook treatment of specific patient groups. The largest area of interest was biomechanics (n=65), followed closely by diabetes (n=61), a third area identified was paediatrics (n=26). Self-perception of specialist status was compared with gender, years of experience, location, primary work environment and clinical practice. When practitioners are asked to categorise themselves to be either “generalist” or “specialist/ generalist with a special interest” podiatrist, male gender was identified as being the only factor which would predict perception of status; 64% males identified as specialist, as opposed to 49% of female survey respondents (Chi square, df = 1, P = 0.044). Self-perception of specialist status was not explained by years of experience, location, working in rural versus urban environment, state worked in, or part-time/full-time work status. Conclusions In conclusion; gender, work environment plus area of interest form a complex relationship, which appear to influence both perception and reality of service provision. Incorporation of specialisation activity (surgical podiatry along with endorsement for use of scheduled medicines) will have lasting impact on the scope of the podiatry profession in Australia. To meet community expectation and maintain high standards, the addition of new subspecialties may be indicated.

Identification of the Sex Pheromone of Holotrichia Reynaudi

The male attractant pheromone of the scarab beetle Holotrichia reynaudi, an agricultural pest native to southern India, was extracted from abdominal glands of females with hexane and analyzed by gas chromatography– mass spectrometry. Field testing of the candidate chemicals, indole, phenol, and anisole, both alone and as binary mixtures, led us to conclude that anisole was the major component of the sex pheromone. Neither male nor female beetles were attracted to indole or phenol on their own. Similarly, when indole and anisole were combined, the attractiveness of the solution did not increase over that obtained with anisole alone. However, combination of phenol and anisole did alter the attractiveness of anisole, with fewer male beetles attracted to the binary mixture than to anisole on its own. The behavior of female beetles was not altered by any of the chemicals tested. Anisole is also the sex pheromone of H. consanguinea, making this the first known example of two melolonthine scarabs sharing the same pheromone.

Identification and preliminary evaluation of viriditoxin, a metabolite of Paecilomyces varioti, as an insecticide for sheep blowfly, Lucilia cuprina (Wied.)

Solvent extracts of cultures of the fungus Paecilomyces varioti are toxic to sheep blowfly, Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae). Different components of the culture extracts were isolated and bioassayed with L. cuprina. The component with most toxicity was purified and identified from its proton magnetic resonance spectrum as viriditoxin, a known antibiotic metabolite of the fungus. The insecticidal properties of viriditoxin were then evaluated. Mean LCso values for first instar larvae of organophosphate susceptible and resistant strains of L. cuprina were 7.5 and 8.4 ppm respectively. Pilot implant trials in sheep demonstrated that the compound provided protection for 9-17 weeks against both strains of L. cuprina. No adverse effects on the trial sheep were detected.

Rapid and efficient screening of a Representational Difference Analysis library using reverse Southern hybridisation: Identification of genetic differences between Haemophilus parasuis isolates

Representational Difference Analysis (RDA) is an established technique used for isolation of specific genetic differences between or within bacterial species. This method was used to investigate the genetic basis of serovar-specificity and the relationship between serovar and virulence in Haemophilus parasuis. An RDA clone library of 96 isolates was constructed using H. parasuis strains H425(P) (serovar 12) and HS1967 (serovar 4). To screen such a large clone library to determine which clones are strain-specific would typically involved separately labelling each clone for use in Southern hybridisation against genomic DNA from each of the strains. In this study, a novel application of reverse Southern hybridisation was used to screen the RDA library: genomic DNA from each strain was labelled and used to probe the library to identify strain-specific clones. This novel approach represents a significant improvement in methodology that is rapid and efficient.

Identification of genes involved with tick infestation in Bos taurus and Bos indicus

Tick resistant cattle could provide a potentially sustainable and environmentally sound method of controlling cattle ticks. Advances in genomics and the availability of the bovine genome sequence open up opportunities to identify useful and selectable genes controlling cattle tick resistance. Using quantitative real-time PCR and the Affymetrix bovine array platform, differences in gene expression of skin biopsies from tick resistant Bos indicus (Brahman) and tick susceptible Bos taurus (Holstein-Friesian) cattle following tick challenge were examined. We identified 138 significant differentially-expressed genes, including several immunological/host defence genes, extracellular matrix proteins, and transcription factors as well as genes involved in lipid metabolism. Three key pathways, represented by genes differentially expressed in resistant Brahmans, were identified; the development of the cell-mediated immune response, structural integrity of the dermis and intracellular Ca 2+ levels. Ca2+, which is implicated in host responses to microbial stimuli, may be required for the enhancement or fine-tuning of transcriptional activation of Ca2+- dependant host defence signalling pathways. Animal Genomics for Animal Health International Symposium, Paris, October 2007: (Proceedings)

Identification and characterization of non-pathogenic Fusarium oxysporum capable of increasing and decreasing Fusarium wilt severity

Fusarium wilt of banana is a potentially devastating disease throughout the world. Options for control of the causal organism, Fusarium oxysporum f.sp. cubense (Foc) are limited. Suppressive soil sites have previously been identified where, despite the presence of Foc, Fusarium wilt does not develop. In order to understand some aspects of this disease suppression, endophytic Fusarium oxysporum isolates were obtained from banana roots. These isolates were genetically characterized and compared with an isolate of Fusarium oxysporum previously identified as being capable of suppressing Fusarium wilt of banana in glasshouse trials. Three additional isolates were selected for glasshouse trials to assess suppression of Fusarium wilt in two different cultivars of banana, Cavendish and Lady Finger. One isolate (BRIP 29089) was identified as a potential biocontrol organism, reducing the disease severity of Fusarium wilt in Lady Finger and Cavendish cultivars. Interestingly, one isolate (BRIP 45952) increased Fusarium wilt disease severity on Cavendish. The implications of an isolate of Fusarium oxysporum, non-pathogenic on banana, increasing disease severity and the potential role of non-pathogenic isolates of Fusarium oxysporum in disease complexes are discussed.

Identification of viral and non-viral reverse transcribing elements in pineapple ( Ananas comosus ), including members of two new badnavirus species

A previously published partial sequence of pineapple bacilliform virus was shown to be from a retrotransposon (family Metaviridae) and not from a badnavirus as previously thought. Two newly discovered sequence groups isolated from pineapple were associated with bacilliform virions and were transmitted by mealybugs. Phylogenetic analyses indicated that they were members of new badnavirus species. A third caulimovirid sequence was also amplified from pineapple, but available evidence suggests that this DNA is not encapsidated, but more likely derived from an endogenous virus.

Pasteurella multocida Expresses Two Lipopolysaccharide Glycoforms Simultaneously, but Only a Single Form Is Required for Virulence: Identification of Two Acceptor-Specific Heptosyl I Transferases

Lipopolysaccharide (LPS) is a critical virulence determinant in Pasteurella multocida and a major antigen responsible for host protective immunity. In other mucosal pathogens, variation in LPS or lipooligosaccharide structure typically occurs in the outer core oligosaccharide regions due to phase variation. P. multocida elaborates a conserved oligosaccharide extension attached to two different, simultaneously expressed inner core structures, one containing a single phosphorylated 3-deoxy-D-manno-octulosonic acid (Kdo) residue and the other containing two Kdo residues. We demonstrate that two heptosyltransferases, HptA and HptB, add the first heptose molecule to the Kdo1 residue and that each exclusively recognizes different acceptor molecules. HptA is specific for the glycoform containing a single, phosphorylated Kdo residue (glycoform A), while HptB is specific for the glycoform containing two Kdo residues (glycoform B). In addition, KdkA was identified as a Kdo kinase, required for phosphorylation of the first Kdo molecule. Importantly, virulence data obtained from infected chickens showed that while wild-type P. multocida expresses both LPS glycoforms in vivo, bacterial mutants that produced only glycoform B were fully virulent, demonstrating for the first time that expression of a single LPS form is sufficient for P. multocida survival in vivo. We conclude that the ability of P. multocida to elaborate alternative inner core LPS structures is due to the simultaneous expression of two different heptosyltransferases that add the first heptose residue to the nascent LPS molecule and to the expression of both a bifunctional Kdo transferase and a Kdo kinase, which results in the initial assembly of two inner core structures.

Identification of QTL for sugar-related traits in a sweet x grain sorghum (Sorghum bicolor L. Moench) recombinant inbred population

QTL for stem sugar-related and other agronomic traits were identified in a converted sweet (R9188) × grain (R9403463-2-1) sorghum population. QTL analyses were conducted using phenotypic data for 11 traits measured in two field experiments and a genetic map comprising 228 SSR and AFLP markers grouped into 16 linkage groups, of which 11 could be assigned to the 10 sorghum chromosomes (SBI-01 to SBI-10). QTL were identified for all traits and were generally co-located to five locations (SBI-01, SBI-03, SBI-05, SBI-06 and SBI-10). QTL alleles from R9188 were detected for increased sucrose content and sugar content on SBI-01, SBI-05 and SBI-06. R9188 also contributed QTL alleles for increased Brix on SBI-05 and SBI-06, and increased sugar content on SBI-03. QTL alleles from R9403463-2-1 were found for increased sucrose content and sucrose yield on SBI-10, and increased glucose content on SBI-07. QTL alleles for increased height, later flowering and greater total dry matter yield were located on SBI-01 of R9403463-2-1, and SBI-06 of R9188. QTL alleles for increased grain yield from both R9403463-2-1 and R9188 were found on SBI-03. As an increase in stem sugars is an important objective in sweet sorghum breeding, the QTL identified in this study could be further investigated for use in marker-assisted selection of sweet sorghum.

Identification of small juvenile scombrids from northwest tropical Australia using mitochondrial DNA cytochrome b sequences

Small juveniles of the nine species of scombrids in Australian waters are morphologically similar to one another and, consequently, difficult to identify to species level. We show that the sequence of the mitochondrial DNA cytochrome b gene region is a powerful tool for identification of these young fish. Using this method, we identified 50 juvenile scombrids collected from Exmouth Bay, Western Australia. Six species of scombrids were apparent in this sample of fish: narrow-barred Spanish mackerel (Scomberomorus commerson), Indian mackerel (Rastrelliger kanagurta), frigate tuna (Auxis thazard), bullet tuna (Auxis rochei), leaping bonito (Cybiosarda elegans), and kawakawa (Euthynnus affinis). The presence of Indian mackerel, frigate tuna, leaping bonito, and kawakawa is the first indication that coastal waters may be an important spawning habitat for these species, although offshore spawning may also occur. The occurrence of small juvenile S. commerson was predicted from the known spawning patterns of that species, but other mackerel species (Scomberomorus munroi, Scomberomorus queenslandicus, Scomberomorus semifasiciatus) likely to be spawning during the sampling period were not detected among the 50 small juveniles analyzed here.

Identification of dynamical systems with fractional derivative damping models using inverse sensitivity analysis

The problem of identifying parameters of time invariant linear dynamical systems with fractional derivative damping models, based on a spatially incomplete set of measured frequency response functions and experimentally determined eigensolutions, is considered. Methods based on inverse sensitivity analysis of damped eigensolutions and frequency response functions are developed. It is shown that the eigensensitivity method requires the development of derivatives of solutions of an asymmetric generalized eigenvalue problem. Both the first and second order inverse sensitivity analyses are considered. The study demonstrates the successful performance of the identification algorithms developed based on synthetic data on one, two and a 33 degrees of freedom vibrating systems with fractional dampers. Limited studies have also been conducted by combining finite element modeling with experimental data on accelerances measured in laboratory conditions on a system consisting of two steel beams rigidly joined together by a rubber hose. The method based on sensitivity of frequency response functions is shown to be more efficient than the eigensensitivity based method in identifying system parameters, especially for large scale systems.

Identification and immunogenicity of an immunodominant mimotope of Avibacterium paragallinarum from a phage display peptide library

Avibacterium paragallinarum is the causative agent of infectious coryza. The protective antigens of this important pathogen have not yet been clearly identified. In this paper, we applied phage display technique to screen the immunodominant mimotopes of a serovar A strain of A. paragallinarum by using a random 12-peptide library, and evaluated the immunogenicity in chickens of the selected mimotope. Polyclonal antibody directed against A. paragallinarum strain 0083 (serovar A) was used as the target antibody and phage clones binding to this target were screened from the 12-mer random peptide library. More than 50% of the phage clones selected in the third round carried the consensus peptide motif sequence A-DP(M)L. The phage clones containing the peptide motif reacted with the target antibody and this interaction could be blocked, in a dose-dependent manner, by A. paragallinarum. One of the peptide sequences, YGLLAVDPLFKP, was selected and the corresponding oligonucleotide sequence was synthesized and then inserted into the expression vector pFliTrx. The recombinant plasmid was transferred into an expression host Escherichia coli GI826 by electroporation, resulting in a recombinant E. coli expressing the peptide on the bacterial surface. Intramuscular injection of the epitope-expressing recombinant bacteria into chickens induced a specific serological response to serovar A. A. paragallinarum. The chickens given the recombinant E. coli showed significant protection against challenge with A. paragallinarum 0083. These results indicated a potential for the use of the mimotope in the development of molecular vaccines for infectious coryza.

Identification of two distinct bovine parainfluenza virus type 3 genotypes

The partial gene sequencing of the matrix (M) protein from seven clinical isolates of bovine parainfluenza virus type 3 (BPIV-3), and the complete sequencing of a representative isolate (Q5592) was completed in this study. Nucleotide sequence analysis was initiated because of the failure of in-house BPIV-3 RT-PCR methods to yield expected products for four of the isolates. Phylogenetic reconstructions based on the nucleotide sequences for the M-protein and the entire genome, using all of the available BPIV-3 nucleotide sequences, demonstrated that there were two distinct BPIV-3 genotypes (BPIV-3a and BPIV-3b). These newly identified genotypes have implications for the development of BPIV-3 molecular detection methods and may also impact on BPIV-3 vaccine formulations.