A novel method for cloning DNA of plant-pathogenic mycoplasmalike organisms

1995 ◽  
Vol 41 (8) ◽  
pp. 753-757 ◽  
Author(s):  
K. H. Chen ◽  
T. A. Chen
Keyword(s):  
Recombinant Dna ◽  
Random Amplified Polymorphic Dna ◽  
Disease Diagnosis ◽  
Oligonucleotide Primer ◽  
Arbitrary Sequence ◽  
Sequence Information ◽  
Dot Blot ◽  
Novel Method ◽  
Recombinant Dna Techniques ◽  
Mycoplasmalike Organisms

A novel method was developed for cloning the DNA from a representative of plant-pathogenic mycoplasmalike organisms (MLOs). This procedure utilized random amplified polymorphic DNA (RAPD) and basic recombinant DNA techniques. It consisted of amplification of total DNA from diseased plants using one oligonucleotide primer with arbitrary sequence and separation of RAPD products in agarose gels. Unique RAPD band(s) of MLO origin was (were) then recovered from the gel and cloned into the specifically designed vector pCRTM II. With this method, a DNA fragment of the SA2 isolate of grapevine yellows MLO was cloned. Southern blot hybridizations revealed that most of the DNA in the unique RAPD band was derived from MLO. Results from dot-blot hybridizations used for screening showed that approximately 60% of transformants harbored MLO-specific recombinant plasmids. Our approach is relatively simple, quite efficient, and not limited by the amount of diseased material available. It does not depend on DNA sequence information for primer design and does not rely on restriction endonucleases for cloning. In addition, it can be used directly for disease diagnosis and for differentiation of closely related MLOs. Our system may serve as a model for cloning DNAs of other fastidious plant pathogens.Key words: mycoplasmalike organism, DNA, cloning, RAPD, detection.

scholarly journals Comparison between the Skin Snip Test and Simple Dot Blot Assay as Potential Rapid Assessment Tools for Onchocerciasis in the Postcontrol Era in Ghana

2002 ◽  
Vol 9 (5) ◽  
pp. 1014-1020 ◽  
Author(s):  
G. E. Guzmán ◽  
K. Awadzi ◽  
N. Opoku ◽  
R. B. Narayanan ◽  
H. O. Akuffo
Keyword(s):  
Recombinant Dna ◽  
Rapid Assessment ◽  
High Specificity ◽  
Diagnostic Methods ◽  
Assessment Tools ◽  
Diagnostic Tools ◽  
Dot Blot ◽  
Native Proteins ◽  
Dot Blot Assay ◽  
Recombinant Dna Techniques

ABSTRACT Successful control of onchocerciasis through mass distribution of ivermectin needs to be coupled with reliable, sensitive, specific, yet affordable diagnostic methods to monitor and ensure the efficacy of such measures. The effort put into the development of diagnostic methods for onchocerciasis that can substitute for or work in combination with the present “gold standard,” the skin snip test, has resulted in the discovery of a number of immunogenic proteins with potential use as diagnostic tools in the postcontrol era. Most of these proteins have now been produced through recombinant DNA techniques. However, when costs are not a trivial issue, none of them have yet found their way into the areas where the disease still exists. In the present study, we have evaluated the performance of a simple dot blot assay which uses a mixture of native proteins designated PakF as a serious contender in the quest for a less invasive and more sensitive method to detect Onchocerca volvulus infection in areas with diverse endemicities. Our results indicate that the assay we propose is more sensitive than the skin snip test and shows high specificity, both characteristics required for a suitable tool for the monitoring of onchocerciasis in the postcontrol era.

Enzymatic Synthesis of 15N-L-aspartic Acid Using Recombinant Aspartase from Escherichia Coli K12

2008 ◽  
Vol 59 (11) ◽  
Author(s):  
Iulia Lupan ◽  
Sergiu Chira ◽  
Maria Chiriac ◽  
Nicolae Palibroda ◽  
Octavian Popescu
Keyword(s):  
Amino Acids ◽  
Aspartic Acid ◽  
Enzymatic Synthesis ◽  
Large Scale ◽  
Recombinant Dna ◽  
Industrial Enzymes ◽  
Recombinant Dna Technology ◽  
Bacterial Fermentation ◽  
Natural Protein ◽  
Novel Method

Amino acids are obtained by bacterial fermentation, extraction from natural protein or enzymatic synthesis from specific substrates. With the introduction of recombinant DNA technology, it has become possible to apply more rational approaches to enzymatic synthesis of amino acids. Aspartase (L-aspartate ammonia-lyase) catalyzes the reversible deamination of L-aspartic acid to yield fumaric acid and ammonia. It is one of the most important industrial enzymes used to produce L-aspartic acid on a large scale. Here we described a novel method for [15N] L-aspartic synthesis from fumarate and ammonia (15NH4Cl) using a recombinant aspartase.

scholarly journals Characterization of an associated microfibril protein through recombinant DNA techniques.

1992 ◽  
Vol 267 (14) ◽  
pp. 10087-10095
Author(s):  
S.K. Horrigan ◽  
C.B. Rich ◽  
B.W. Streeten ◽  
Z.Y. Li ◽  
J.A. Foster
Keyword(s):  
Recombinant Dna ◽  
Recombinant Dna Techniques

Report of the committee on human gene mapping by recombinant DNA techniques

1984 ◽  
Vol 37 (1-4) ◽  
pp. 210-273 ◽  
Author(s):  
M.H. Skolnick ◽  
H.F. Willard ◽  
L.A. Menlove
Keyword(s):  
Gene Mapping ◽  
Recombinant Dna ◽  
Human Gene ◽  
Human Gene Mapping ◽  
Recombinant Dna Techniques

Recombinant DNA techniques; an introduction

Gene ◽  
1983 ◽  
Vol 26 (2-3) ◽  
pp. 323-324
Author(s):  
A.J. Podhajska
Keyword(s):  
Recombinant Dna ◽  
Recombinant Dna Techniques

Recombinant DNA techniques in diagnostic and preventive medicine

BioEssays ◽  
1984 ◽  
Vol 1 (1) ◽  
pp. 12-15 ◽  
Author(s):  
Stephen Hodgkinson ◽  
Peter Scambler
Keyword(s):  
Preventive Medicine ◽  
Recombinant Dna ◽  
Recombinant Dna Techniques

scholarly journals Influenza virus hemagglutinin-specific cytotoxic T cell response induced by polypeptide produced in Escherichia coli.

1985 ◽  
Vol 162 (2) ◽  
pp. 663-674 ◽  
Author(s):  
A Yamada ◽  
M R Ziese ◽  
J F Young ◽  
Y K Yamada ◽  
F A Ennis
Keyword(s):  
Influenza Virus ◽  
Recombinant Dna ◽  
Cytotoxic T Lymphocyte ◽  
Nonstructural Protein ◽  
T Cell Response ◽  
Cytotoxic T Cell ◽  
Protein Immunization ◽  
Recombinant Dna Techniques

We have tested the abilities of various polypeptides of A/PR/8/34 (H1N1) virus, constructed by recombinant DNA techniques, to induce influenza virus-specific secondary cytotoxic T lymphocyte (CTL) responses. A hybrid protein (c13 protein), consisting of the first 81 amino acids of viral nonstructural protein (NS1) and the HA2 subunit of viral hemagglutinin (HA), induced H-2-restricted, influenza virus subtype-specific secondary CTL in vitro, although other peptides did not. Using a recombinant virus, the viral determinant responsible for recognition was mapped to the HA2 portion of c13 protein. Immunization of mice with c13 protein induced the generation of memory CTL in vivo. The CTL precursor frequencies of A/PR/8/34 virus- and c13 protein-immune mice were estimated as one in 8,047 and 50,312, respectively. These results indicate that c13 protein primed recipient mice, even though the level of precursor frequency was below that observed in virus-immune mice.

scholarly journals Molecular biology of baculovirus and its use in biological control in Brazil

1999 ◽  
Vol 34 (10) ◽  
pp. 1733-1761 ◽  
Author(s):  
Maria Elita Batista de Castro ◽  
Marlinda Lobo de Souza ◽  
William Sihler ◽  
Júlio Carlyle Macedo Rodrigues ◽  
Bergmann Morais Ribeiro
Keyword(s):  
Molecular Biology ◽  
Recombinant Dna ◽  
Host Cells ◽  
Virus Propagation ◽  
Viral Particles ◽  
Occlusion Bodies ◽  
Budded Virus ◽  
Recombinant Dna Techniques ◽  
High Level ◽  
Eukaryotic Organisms

Baculoviruses are insect viruses found mainly in Lepidoptera. The family Baculoviridae is taxonomically divided in two genera, Nucleopolyhedrovirus and Granulovirus, which differ by occlusion body morphology. NPVs (Nucleopolyhedroviruses) have polyhedrical inclusion bodies (PIBs) containing multiple viral particles, while GVs (Granuloviruses) appear to be generally single particles occluded in oval shaped occlusion bodies. During the life cycle, two different viral progenies are produced: BV (Budded Virus) and PDV (Polyhedra Derived Virus), which are essential for the infectious process and virus propagation in host cells. Baculoviruses are being used for pest control and they are especially safe due to their specificity and invertebrate-restricted host range. Baculoviruses have been used as vectors for high level protein expression ofheterologous genes from prokaryotic and eukaryotic organisms. Also, recombinant DNA techniques have allowed the production of genetically modified viral insecticides. This study is a review on the taxonomy, structure, replication and molecular biology of baculoviruses, as well as their use as bioinsecticides in Brazil.

The Growing Role of Genetic Professionals in Forensic DNA

2009 ◽  
Vol 6 (5) ◽  
Author(s):  
Cristina Pinto
Keyword(s):  
Forensic Medicine ◽  
Tandem Repeats ◽  
Nuclear Dna ◽  
Recombinant Dna ◽  
Dna Typing ◽  
Individual Identification ◽  
Bar Code ◽  
Genomic Polymorphism ◽  
Recombinant Dna Techniques ◽  
Dna Profiles

AbstractThrough the last decade there was an enormous revolution in the field of forensic genetic.The Author reviews some of the methodologies used in the definitions of DNA profiling tackling the principles of recombinant DNA techniques. The potentiality of polymorphic DNA fragments in vertebrates is focused as well as the revolution implied in forensic medicine. The resource to DNA-DNA hybridization combined to oligonucleotide probes is emphasized leading to the production of an individual bar code with the resource of genomic polymorphism which leads to a pattern known as genetic fingerprinting. Other techniques for individual identification and paternity testing are focused as well as the use of short tandem repeats (STR's). Mitochondrial DNA sequencing use to complement nuclear DNA typing may also be profitable in certain instances. Relevant problems within the context of the use of these techniques in forensic medicine and law suits are discussed. Final considerations viewing the resource to DNA technology within the scope of the last two decades are referred regarding the resource to DNA profiles not only in the US but in Europe in general and in Portugal in special having lead to compensation and uncover of justice errors.

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