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KMID : 1007519970060010030
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Food Science and Biotechnology
1997 Volume.6 No. 1 p.30 ~ p.33
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Optimization of PCR for RAPD Analysis of Bifidobacterium spp .
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SO, JAE SEONG
HEO, TAE RYEON/Song, Ji Eun
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Abstract
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Random amplified polymorphic DNA (RAPD) is based on the use of a single arbitrary primer in the polymerase chain reaction (PCR) to amplify segments of the genome which could indicate the polymorphism of the sample DNA. The aim of this study was to establish simple and rapid identification procedures for Bifidobacterium spp. by RAPD technique. The optimal condition for PCR was obtained in a solution with a total volume of 50 ㎕ containing 20 ng of template DNA, 0.8 μM random primer(5´GTAACGCC3´), 2.0 U Tag polymerase, 2.5 mM MgCl₂and 100 μM dNTPs. Samples were amplified for 45 cycles of 1 min at 94℃, 1 min at 35℃ and 1 min at 72℃. Under this condition we observed different band patterns of resulting PCR products on agarose gels, which could distinguish among different Bifidobacterium spp. Furthermore, for rapid RAPD analysis we have attempted to use DNA from freeze-thaw lysed cells without purification of the DNA sample, and successful amplifications were also obtained. This latter method took only 7 hours to complete RAPD analysis from DNA preparation to electrophoresis.
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KEYWORD
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