Comparative evaluation of three different extraction methods for rice (Oryza sativa L.) genomic DNA.
Recently, more immediate and precise cultivar-identifying methods targeting the specific and/or introduced gene(s) have been put into practical use for various rice cultivars. However trustworthy and innovative the biotechnological analyses may be, DNA purity and quality do have unpredictable influences upon the identification. Extraction methods of rice DNA have hardly ever been compared in a comprehensive manner. In this study, we investigated extraction characteristics of three methods by using 10 rice cultivars and then examined template availability of rice DNAs thereby extracted. An UV spectrophotometric study with a view toward methods revealed three different facts: The Isoplant II kit method with inhibitor absorption yielded the most DNAs, the Takara kit method with magnetic trapping produced the best DNAs free from contaminative proteins, and the enzymatic digestion method exclusively with enzymatic digestions prepared the best DNAs free from contaminative polysaccharides. Moreover, with a view toward cultivars, an insignificant difference in yield was not entirely bore out, and some difference in cultivar might cause significant difference in yield; however, no significant difference in purity was found among the cultivars used. On the other hand, electrophoretic images of the DNAs from the same cultivars showed considerable differences in quality among the methods. Furthermore, the DNA extracts from certain brands of rice proved really available for cultivar identification by using polymerase chain reaction (PCR) related to sequence-tagged sites. Therefore, this study suggested that these extraction methods may be used as the situation demands and that the DNAs thereby extracted might work successfully even in cultivar-identifying PCRs.
Sagi
Naoki
N
Tokyo Metropolitan Institute of Public Health, 24-1 Hyakunincho-3, Shinjuku-ku, Tokyo 169-0073, Japan. Naoki_Sagi@member.metro.tokyo.jp
Monma
Kimio
K
Ibe
Akihiro
A
Kamata
Kunihiro
K
D018744:DNA, Plant / Q000302:isolation & purification*; D004587:Electrophoresis, Agar Gel; D007564:Japan; D012275:Oryza / Q000145:classification / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011933:Reagent Kits, Diagnostic; D012639:Seeds / Q000235:genetics; D013053:Spectrophotometry; D000516:alpha-Amylases
D018744:DNA, Plant; D011933:Reagent Kits, Diagnostic; D000516:alpha-Amylases