ISSN:
1573-0972
Keywords:
Arbitrary primers
;
Brucella abortus
;
Brucella melitensis
;
polymerase chain reaction
;
RAPD
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
,
Process Engineering, Biotechnology, Nutrition Technology
Notes:
Abstract Randomly amplified polymorphic DNA (RAPD) profiles of Brucella and non-Brucella DNA were established after polymerase chain reaction (PCR) amplification. Five arbitrary oligonucleotide primers were screened to generate Brucella-specific DNA fingerprints. The arbitrary primer OPB-01 (5′-GTTTCGCTCC-3′) produced DNA bands specific to Brucella. Amplification conditions must be optimized for reproductibility. Accordingly, we optimized and established the conditions, which included Mg2+, enzyme (DNA polymerase), primer, template and deoxyribonucleoside triphosphate (dNTP) concentrations as well as the optimum number of thermal cycles to produce OPB-01 directed Brucella DNA fingerprints. The optimized RAPD method can produce a 1.3 kb DNA fragment specific to Brucella. This DNA fragment was common to eight biovars of B. abortus and one biovar of B. melitensis. The fragment was not detected in genetically related species such as Ochrobactrum anthropi and other non-Brucella organisms associated with farm animals. We anticipate the use of this fragment as a possible probe for the detection of Brucella organisms.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1023/A:1008877630986
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