학술논문
應用單一步驟巢式複合式聚合酶鏈鎖反應同步鑑定刺足根蟎、羅賓根蟎及長毛根蟎 / The application of single-step nested multiplex polymerase chain reaction for the identification of Rhizoglyphus echinopus, R. robini and R. setosus simultaneously
Document Type
Article
Author
Source
植物保護學會會刊. Vol. 48 Issue 2, p101-116. 16 p.
Subject
Language
繁體中文
ISSN
0577-750X
Abstract
A specific single-step nested polymerase chain reaction (PCR) assay was developed to identify the three species of worldwide economically important bulb mites: Rhizoglyphus echinopus, R. robini and R. setosus simultaneously. General primers were used to amplify the 18S-28S region of ribosomal DNA by PCR. Sequences of the ITS1 region of the PCR products were used to construct species-specific primers which were then used to amplify the species specific DNA fragments. These fragments which can be visualized readily after electrophoresis were 522, 312 and 404 bp for R. echinopus, R. robini and R. setosus, respectively. The plus strand universal primers derived from the sequence in the 18S and 5.8S coding region increase the sensitivity of this single-step nested multiplex PCR assay to pg degree. Treatment of these species-specific DNA fragment with Taq I, Mfe I, Sty I and Ssp I further increase the accuracy in differencing these three species.