Selection of Polymerase Chain Reaction Primers from an RNA Intergenic Spacer Region for Specific Detection ofClavibacter michiganensissubsp.sepedonicus

1995 ◽  
Vol 85 (8) ◽  
pp. 837 ◽  
Author(s):  
Xiang Li
Keyword(s):  
Polymerase Chain Reaction ◽  
Intergenic Spacer ◽  
Specific Detection ◽  
Chain Reaction ◽  
Intergenic Spacer Region ◽  
Polymerase Chain ◽  
Polymerase Chain Reaction Primers ◽  
Selection Of

scholarly journals Differentiation of Moraxella Bovoculi sp. nov. from other Coccoid Moraxellae by the Use of Polymerase Chain Reaction and Restriction Endonuclease Analysis of Amplified DNA

2007 ◽  
Vol 19 (5) ◽  
pp. 532-534 ◽  
Author(s):  
John A. Angelos ◽  
Louise M. Ball
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Sequence ◽  
Restriction Analysis ◽  
Intergenic Spacer ◽  
Restriction Enzyme Digestion ◽  
Chain Reaction ◽  
Gram Negative ◽  
Intergenic Spacer Region ◽  
Polymerase Chain ◽  
Moraxella Bovoculi

Moraxella oris was historically the only coccoid Moraxella identified in cultures of ocular fluid from cattle with infectious bovine keratoconjunctivitis (IBK) and could be morphologically and biochemically differentiated from Moraxella bovis. Moraxella bovoculi sp. nov. is a recently characterized Moraxella isolated from ulcerated eyes of calves with IBK in northern California in 2002. Like Moraxella ovis, M. bovoculi sp. nov. is a gram-negative coccus/diplococcus. All 18 original isolates of M. bovoculi sp. nov. possessed phenylalanine deaminase (PADase) activity and could therefore be differentiated from M. ovis and M. bovis. During the characterization of 44 additional isolates of hemolytic gram-negative cocci that were cultured from ulcerated eyes of IBK-affected calves, 2 PADase-negative isolates were identified that could not be differentiated biochemically from M. ovis; however, the DNA sequence of the 16S-23S intergenic spacer region (ISR) of the isolates matched the 16S-23S ISR DNA sequence of M. bovoculi sp. nov. To facilitate the identification of PADase-negative moraxellae, a polymerase chain reaction (PCR) coupled with restriction enzyme digestion analysis of amplified DNA was developed. Amplification of the 16S-23S ISR followed by AfaI digestion of amplified DNA could differentiate M. bovoculi sp. nov. from M. ovis and other moraxellae. The DNA sequence analysis of the amplified 16S-23S ISR from the 42 PADase-positive isolates of hemolytic gram-negative cocci indicated that all were M. bovoculi sp. nov. and all possessed an AfaI site. A PCR coupled with restriction analysis of amplified DNA can aid in identifying M. bovoculi sp. nov.

Rapid Polymerase Chain Reaction/DNA Probe Membrane-Based Assay for the Detection of Listeria and Listeria monocytogenes in Food

2000 ◽  
Vol 63 (3) ◽  
pp. 337-342 ◽  
Author(s):  
L. O'CONNOR ◽  
J. JOY ◽  
M. KANE ◽  
T. SMITH ◽  
M. MAHER
Keyword(s):  
Polymerase Chain Reaction ◽  
Microbiological Quality ◽  
Intergenic Spacer ◽  
Pcr Primers ◽  
Dna Probe ◽  
23S Rrna ◽  
Chain Reaction ◽  
Intergenic Spacer Region ◽  
Rapid Polymerase Chain Reaction ◽  
Polymerase Chain

We describe the development of polymerase chain reaction (PCR)/DNA probe membrane-based colorimetric assays for the detection and identification of Listeria and L. monocytogenes. PCR primers designed from the 16S to 23S rRNA intergenic spacer region amplified products that were reverse hybridized to membrane-bound oligonucleotide probes specific for Listeria and L. monocytogenes with a detection limit of 1 to 10 CFU/25 ml in inoculated raw and pasteurized milk samples. These qualitative assays have the potential to be integrated into testing laboratories for monitoring the microbiological quality of foods.

Genus- and species-specific detection ofListeria monocytogenesusing polymerase chain reaction assays targeting the 16S/23S intergenic spacer region of the rRNA operon

1996 ◽  
Vol 42 (11) ◽  
pp. 1155-1162 ◽  
Author(s):  
Tom Graham ◽  
Elizabeth J. Golsteyn-Thomas ◽  
Victor P. J. Gannon ◽  
James E. Thomas
Keyword(s):  
Polymerase Chain Reaction ◽  
Listeria Monocytogenes ◽  
Sequence Data ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Chain Reaction ◽  
Nucleotide Sequence Data ◽  
Intergenic Spacer Region ◽  
Polymerase Chain ◽  
Species Specific

In this study, the 16S/23S rRNA intergenic spacer (IGS) regions of six Listeria species were examined. DNA bands of 590 and 340 bp were observed following polymerase chain reaction (PCR) amplification of DNA from Listeria monocytogenes, Listeria innocua, Listeria seeligeri, Listeria welshimeri, and Listeria ivanovii strains with generic rRNA IGS oligonucleotide primers. For strains of Listeria grayi subspp. grayi and murrayi, DNA band sizes of 550 and 340 bp were observed with this primer pair. DNA bands of these sizes were not observed for other Gram-negative or -positive bacteria in this PCR assay. Four RsaI digestion profiles were noted for the Listeria PCR products. Listeria monocytogenes strains had one profile; L. innocua strains had a second; L. seeligeri, L. welshimeri, and L. ivanovii strains had a third; and L. grayi strains had a fourth. The small and large 16S/23S rRNA IGSs of L. monocytogenes ATCC 15313 were identical in the first 58 5′ and the last 169 3′ nucleotides. However, the large rRNA IGS contained a central 267-bp region with tRNAIleand tRNAAlagenes. Large rRNA 16S/23S IGS nucleotide sequence data has not been previously reported. This data was used to develop novel Listeria genus-specific and L. monocytogenes species-specific PCR assays.Key words: polymerase chain reaction, 16S/23S rRNA intergenic spacer region, Listeria monocytogenes.

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