Identification of Clavibacter michiganensis subsp. sepedonicus using the polymerase chain reaction

1994 ◽  
Vol 40 (2) ◽  
pp. 148-151 ◽  
Author(s):  
Giuseppe Firrao ◽  
Romano Locci
Keyword(s):  
Polymerase Chain Reaction ◽  
Clavibacter Michiganensis ◽  
Chain Reaction ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Clavibacter Michiganensis Subsp ◽  
Ring Rot ◽  
Dna Fragment ◽  
Polymerase Chain ◽  
Polymerase Chain Reaction Primers

From the sequence of a subcloned DNA fragment of a highly conserved plasmid of Clavibacter michiganensis subsp. sepedonicus a pair of oligonucleotides were devised for use as polymerase chain reaction primers. The primer sequences do not show significant homology with any other sequence deposited in public databases. Polymerase chain reactions carried out using this primer pair and untreated cells of all strains of C. michiganensis sepedonicus tested resulted in the amplification of a DNA fragment of about 670 base pairs. No amplification was observed when bacteria belonging to other species were submitted to polymerase chain reaction under the same conditions. The detection limit of the assay was 4 × 103 bacteria.Key words: detection, potato ring rot, Corynebacterium sepedonicum.

Investigation of Mutations in Exon 12 Of MYH7, 16 Of β-MYBPC3 and 2 Of TCAP Gene in Dogs with Dilated Cardiomyopathy using PCR-SSCP Technique

2021 ◽  
Author(s):  
R.B. Vishnurahav ◽  
S. Ajithkumar ◽  
Usha Narayana Pillai ◽  
N. Madhvan Unny ◽  
K.D. John Martin ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Dilated Cardiomyopathy ◽  
Genetic Alterations ◽  
Chain Reaction ◽  
Multifunctional Protein ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Intron 1 ◽  
Cardiac Disorders ◽  
Polymerase Chain

Background: Dilated cardiomyopathy is the important myocardial disease and one of the most common cause of death in the medium to large size dog breeds worldwide. The disease is characterized by dilatation of cardiac chambers and thinning of walls leads to systolic failure. Mutations in some sarcomere genes leads to cardiomyopathy in humans. Sarcomere is an important multifunctional protein network involved in the signal reception and transduction. Mutations in β-MYH7, MYBPC3 and TCAP genes produce alterations in the morphology of heart (hypertrophy or dilatation).Methods: In this study twenty apparently healthy and twenty five dogs with dilated cardiomyopathy (DCM) were selected from patients reported or referred to University Veterinary Hospital and Teaching Veterinary Clinical Complex, Mannuthy (2015-2017) based on the clinical examination, radiographic, electrocardiographic, haematobiochemical and echocardiographic studies cardiac disorders (Dilated cardiomyopathy and hypertrophic cardiomyopathy) were confirmed.Result: In the present study we investigated genetic alterations of exon 12 of MYH7, 16 of β-MYBPC3 and 2 of TCAP gene in dogs by polymerase chain reaction -single stranded confirmation of polymorphism (PCR-SSCP). Polymerase chain reactions were analysed using acrylamide gel and samples with different pattern of bands were sequenced. Polymerase chain reaction-SSCP showed different migration of band pattern in the intron 1 of TCAP gene in one sample.

Detection and identification of Clavibacter michiganensis subsp. sepedonicus and Clavibacter michiganensis subsp. michiganensis by nonradioactive hybridization, polymerase chain reaction, and restriction enzyme analysis

1994 ◽  
Vol 40 (12) ◽  
pp. 1007-1018 ◽  
Author(s):  
J. L. W. Rademaker ◽  
J. D. Janse
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Product ◽  
Enzyme Analysis ◽  
Restriction Enzyme Analysis ◽  
Bacterial Strains ◽  
Clavibacter Michiganensis ◽  
Chain Reaction ◽  
Clavibacter Michiganensis Subsp ◽  
Detection And Identification ◽  
Polymerase Chain

To develop a rapid and reliable detection and identification method for Clavibacter michiganensis subsp. sepedonicus and C. michiganensis subsp. michiganensis, two biotinylated probes and derived primer sets were evaluated for specificity using a large number of bacterial strains. Detection in dot blot analysis using the Diagen probe against C. michiganensis subsp. sepedonicus was possible with all 32 C. michiganensis subsp. sepedonicus strains tested. Cross-hybridization occurred with all nine C. michiganensis subsp. insidiosus strains tested. No hybridization occurred with any of 54 other related and unrelated bacterial strains including C. michiganensis subsp. michiganensis, C. michiganensis subsp. nebraskensis, C. michiganensis subsp. tessellarius, C. iranicus, C. rathayi, and C. tritici and potato saprophytes. Hybridization of the MIC 1 probe against C. michiganensis subsp. michiganensis was obtained with 22 out of 24 C. michiganensis subsp. michiganensis strains. A weak hybridization signal occurred only with two strains of C. michiganensis subsp. insidiosns. No hybridization occurred with any of the 71 other related and unrelated bacterial strains tested including tomato saprophytes. Restriction fragment length polymorphisms detected with the Diagen probe allowed differentiation between C. michiganensis subsp. sepedonicus and the related C. michiganensis subsp. insidiosus. Restriction fragment length polymorphism analysis using the MIC 1 probe and BamH1 showed at least two groups of patterns within C. michiganensis subsp. michiganensis. By using a primer set derived from the Diagen probe, a DNA sequence could be amplified with all C. michiganensis subsp. sepedonicus strains tested. Only the nontarget organism C. michiganensis subsp. insidiosus yielded a similar polymerase chain reaction product. Restriction enzyme analysis of the polymerase chain reaction product enabled rapid distinction between the subspecies. With a CMM primer set derived from the MIC 1 probe a DNA sequence was amplified from the same 22 out of 24 C. michiganensis subsp. michiganensis strains that showed hybridization with the MIC 1 probe. The polymerase chain reaction product could be verified by restriction enzyme analysis. The Diagen and MIC 1 probes and derived primer sets were shown to be useful for the detection and identification of C. michiganensis subsp. sepedonicus and C. michiganensis subsp. michiganensis. The MIC 1 probe, however, failed to detect two strains of the latter subspecies.Key words: biotin, PCR, REA, potato bacterial ring rot, bacterial canker of tomato, RFLP, Clavibacter michiganensis subsp. insidiosus.

Detection of rye chromosome 2R using the polymerase chain reaction and sequence-specific DNA primers

Genome ◽  
1994 ◽  
Vol 37 (1) ◽  
pp. 19-22 ◽  
Author(s):  
J.-H. Lee ◽  
R. A. Graybosch ◽  
D. J. Lee
Keyword(s):  
Polymerase Chain Reaction ◽  
Wheat Chromosome ◽  
Specific Marker ◽  
Chain Reaction ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Pcr Product ◽  
Polymerase Chain ◽  
Wheat Lines ◽  
Dna Primers

Sequences derived from a rye gamma secalin gene were used as primers in polymerase chain reactions using DNA obtained from a series of wheat and triticale genetic stocks. A 473-bp fragment, the predicted size based on the distance between the selected primers, was found only in rye, triticales, and wheat lines carrying rye chromosome 2RS. Use of triticale lines with various wheat chromosome substitutions confirmed the chromosomal origin of the rye-specific marker. The presence of the 473-bp PCR product was always associated with the production of 75K secalins in grain samples. Thus, the primer sequences, and the clone of origin (pSC503), were both derived from the SEC-2 locus of rye chromosome 2RS.Key words: wheat (Triticum aestivum), rye (Secale cereale), chromosomal translocations, chromosomal substitutions, DNA polymerase chain reaction, sequence-specific primers.

scholarly journals Detection of Bovine Leukemia Virus in Blood and Milk by Nested and Real-Time Polymerase Chain Reactions

2003 ◽  
Vol 15 (1) ◽  
pp. 72-76 ◽  
Author(s):  
Christopher J. Kuckleburg ◽  
Christopher C. Chase ◽  
Eric A. Nelson ◽  
Salvatore A. E. Marras ◽  
Matthew A. Dammen ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Early Detection ◽  
Real Time ◽  
Bovine Leukemia Virus ◽  
Leukemia Virus ◽  
Chain Reaction ◽  
Nested Polymerase Chain Reaction ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Polymerase Chain

Concerns about retroviruses in livestock and products derived from them have necessitated the development of tests to detect the bovine leukemia virus (BLV) in blood and milk from cattle. Dairy cattle ( n = 101) from 5 different geographical areas were used for this study. A nested polymerase chain reaction (PCR) identified 98% of BLV seropositive cattle ( n = 80) from blood and 65% from milk, whereas real-time PCR detected 94% of BLV seropositive cattle from blood and 59% from milk. Bovine leukemia virus was also detected by PCR in approximately 10% of seronegative cattle ( n = 21), most likely because of early detection before seroconversion.

scholarly journals A Guide to Cloning the Products of Polymerase Chain Reactions

2021 ◽  
Vol 2021 (9) ◽  
pp. pdb.top101345
Author(s):  
Michael R. Green ◽  
Joseph Sambrook
Keyword(s):  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Polymerase Chain

This introduction outlines various methods to clone amplified DNAs and to facilitate the construction of complex multicomponent genetic units. Because of the ease with which the termini of amplified DNAs can be tailored by polymerase chain reaction (PCR), many of the methods outlined here use PCR not only to synthesize DNAs but also to link them together into purpose-designed constructs. The most recent refinements however have been the development of modular genetic units that can be harnessed to target DNAs not by PCR but by site-specific recombination enzymes.

scholarly journals Testing the effect of new biopesticides on soilborne pathogens of vegetable diseases using DNA markers

2020 ◽  
Vol 10 (3) ◽  
pp. 401-411
Author(s):  
N. E. Pavlovskaya ◽  
I. N. Gagarina ◽  
A. Yu. Gavrilova ◽  
D. B. Borodin
Keyword(s):  
Polymerase Chain Reaction ◽  
Vegetable Crops ◽  
Clavibacter Michiganensis ◽  
Chain Reaction ◽  
Fungicidal Action ◽  
Clavibacter Michiganensis Subsp ◽  
Polymerase Chain ◽  
Xanthomonas Euvesicatoria ◽  
Biological Protection ◽  
Biological Methods

In protected ground, vegetables are exposed to various pests and pathogens of viral, bacterial or fungal diseases. Growing vegetables indoors requires the elimination of all chemical treatments and control over pathogens and pests. In this regard, the application of sensitive methods based on molecular markers in the diagnostics of pathogens is highly relevant. In this work, polymerase chain reaction was used to identify the most common diseases of tomato (Xanthomonas euvesicatoria), cucumber (Ascochyta cucumis) and potato (Clavibacter michiganensis subsp. Sepedonicus) plants. It was established that early diagnosis of vegetable diseases using polymerase chain reaction allows rapid detection of trace amounts of pathogenic microorganisms thus facilitating selection of effective fungicidal preparations. The creation of biological protection for vegetable crops is relevant considering their nutritional importance. New biological methods for protecting vegetables in greenhouses were tested. Thus, the fungicidal effect of the authors’ patented preparation developed for pre-sowing treatment of vegetable seeds in protected ground on the pathogens of tomato  black  bacterial  spot  (Xanthomonas  euvesicatoria)  and  cucumber  stem  hypertrophy (Ascochyta cucumis) was confirmed. Another preparation patented as a means of pre-sowing treatment of pea seeds demonstrated fungicidal action against the development of Phytophthora infectants on potato plants. It was observed that these preparations are effective at concentrations of 10-4 % for soaking seeds and double treatment of plants during the flowering period.

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